Difference between revisions of "Team:Freiburg/NotebookTargeting"

Latest revision as of 23:20, 19 October 2016

Lab Journals

Group 4 - Adhesion

The targeting is one of the main requirements the spores have to fulfill. An adhesion-assay was conducted to verify the binding of the spores to a desired target.

07/04/2016

Bacillus subtilis strains used:

Strain

B52:B. Subtilis w168 amyE::PcgeA-CotZre-gfp-B0014(CM5)

B53:B. Subtilis w168 amyE::PcotYZ-CotZre-gfp-B0014(CM5)

B54:B. Subtilis w168 amyE::PcotYZ-CotZin-gfp-B0014(CM5)

B55:B. Subtilis w168 amyE::PcotV-CotZre-gfp-B0014(CM5)

B56:B. Subtilis w168 amyE::PCotv-CotZin-gfp-B0014(CM5)

07/05/2016

Slide preparation: GOPTS

Surface activation of glass slides by plasma generator, coated with GOPTS 3-glycidyloxypropyl trimethoxysilane, washed with acetone and then stored for up to two weeeks.

labeled with a diamond pen
washed with EtOH then ddH2O
dried slides with compressed air
activation of surface with plasma generator
incubation with GOPTS (overnight)

07/10/2016

Buffer for anti-GFP nanobody (10 ml)

61mg Tris
87mg NaCl
1ml Glycerin
2,5 ml Imidazole(1M)
fill up with ddH2O to 10 ml

07/12/2016

Sample preparation 1: GOPTS + anti-GFP nanobody + B52-B56

incubation with nanobody (0.5µl, 330µg/ml, overnight, 4°C)

07/13/2016

Sample preparation 1: GOPTS + anti-GFP nanobody + B52-B56

blocked with BSA(2%) (150µl/spot, 30 min)
washed with PBS(1x)
washed with imidazole (20mM) in PBS(1x)
washed with PBS(1x)
washed with ddH2O
dried with compressed air
incubation with B52-B56 (100µl/spot, 120 min)
incubation with Mowiol DABCO(1,4-Diazabicyclo[2.2.2]octane) (5µl/spot, overnight), with second glass slide on top

Spot No.	Sample
1	GOPTS + nanobody + B52
2	GOPTS + nanobody + B53
3	GOPTS + nanobody + B54
4	GOPTS + nanobody + B55
5	GOPTS + nanobody + B56

07/14/2016

Sample preparation 1: GOPTS + anti-GFP nanobody + B52-B56

edges sealed with nail polish

Fluorescence microscopy 1 (Olympus 1×71): GOPTS + anti-GFP nanobody + B52-B56

–> pictures inconclusive: wrong coverslips (second glass slide), no sufficient controls

Sample preparation 2: GOPTS + anti-GFP nanobody + B52-56

incubation with anti-GFP nanobody (5µl/spot, 330 µg/ml, 120 min)
blocked with BSA(2%) (90 min)
rinsed with PBS(1x)
stored in PBS(1x) (for 9 days)

Spot No.	Sample
1	GOPTS + nanobody + B52
2	GOPTS + nanobody + B53
3	GOPTS + nanobody + B54
4	GOPTS + nanobody + B55
5	GOPTS + nanobody + B56
6	GOPTS + GFP (positive control)
7	GOPTS + WT (negative control)

07/15/2016

Sample preparation

added nanobody to GOPTS plate (300 µg/ml, 5µl)
tried to pipette rest nanobody down, but didn´t work
blocked complete slide with BSA(2%)

07/23/016

Sample preparation 2: GOPTS + anti-GFP nanobody + B52-56

washed slides with ddH2O
incubation with B52, B53, B54, B55, B56 (50µl/spot, overnight)

–> FACS-analysis has shown that only B53 and B54 show fluorescence linked to GFP

07/24/2016

Preparation of samples 3: B53-B55

B53, B54, B55 (10µl/spot) on uncoated glass slide
Mowiol DABCO (5µl/spot)
covered with cover slip

Spot No.	Sample
1	B52
2	B53
3	B54

Fluorescence microscopy 2 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + B52-56

–> fluorescence signal of B53 and B54

Fluorescence microscopy 3 (Olympus 1×71): B53-B55

–> fluorescence signal of positive controls (B53 and B54) but no signal of spores bound to GOPTS

Sample preparation 4: GOPTS + anti-GFP nanobody + B53/B54

incubation with anti-GFP nanobody (5µl, 330µg/ml, 2d)

07/26/2016

Sample preparation 4: GOPTS + anti-GFP nanobody + B53/B54

blocked with BSA(4%) (overnight)

07/27/2016

Sample preparation 4: GOPTS + anti-GFP nanobody + B53/B54

rinsed and washed with PBS(1x) (10 min, 10rpm)
rinsed with water ddH2O
dried with compressed air
incubation with Mowiol DABCO (5µl/spot, overnight)

Spot No.	Sample
1	GOPTS + GFP (positive control)
2	GOPTS + nanobody + GFP
3	GOPTS + BSA + GFP (negative control)
4	GOPTS + B53 (positive control)
5	GOPTS + B54 (positive control)
6	B53 (positive control)
7	B54 (positive control)

07/28/2016

Fluorescence microscopy 4 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + B53/B54

–> spores appear to not have been bound to nanobody

08/08/2016

Sample preparation 5: GOPTS + anti-GFP nanobody + B53-56/GFP

incubation with anti-GFP nanobody (5µl, 330µg/ml, 60 min)
rinsed and blocked with BSA(4%) (30 min)
incubation with B53, B54, B55, B56, GFP (5µl/spot, 60 min)
rinsed and washed with PBS(1x) (10 min, 10 rpm)
rinsed with water ddH2O
dried with compressed air
incubation with Mowiol DABCO (5µl/spot, overnight)

Spot No.	Sample
1	GOPTS + nanobody + B55 (negative control)
2	GOPTS + B55 (negative control)
3	GOPTS + nanobody + B53
4	GOPTS + nanobody + GFP (positive control)
5	GOPTS + BSA + GFP (negative control)
6	B53 (positive control)
7	B54 (positive control)
8	GOPTS + GFP (positive control)
9	GOPTS + B53 (positive control)
10	GOPTS + nanobody(Alexa647) + B53
11	GOPTS + nanobody(Alexa647) + B54
12	GOPTS + nanobody(Alexa647) + WT(negative control)
13	GOPTS + nanobody(Alexa647) + B56 (negative control)

08/09/2016

Fluorescence microscopy 5 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + B53-56/GFP

–> spores have not been bound

08/10/2016

Sample preparation 6: anti-GFP nanobody + spores (WT, B53, B54, B56) (in solution)

incubation of anti-GFP nanobody (Alexa647-conjugated) (1µl, 330µg/ml, 60 min) + spores (WT, B53, B54, B56) (400µl, 60 min, 4°C)
centrifuged (13400 rpm, 2 min)
washed with PBS(1x) (1 ml)
repeated steps 2 and 3
centrifuged (13400 rpm, 2 min)
resuspended in PBS(1x) (400µl)

Spot No.	Sample
1	nanobody + WT (negative control)
2	nanobody + B53
3	nanobody + B53
4	nanobody + B55 (negative control)
5	nanobody + B56(negative control)
6	nanobody + GFP (positive control)
7	nanobody (positive control)
8	GFP (positive control)

Fluorescence microscopy 6 (Nikon C2+ confocal microscope): anti-GFP nanobody + spores (WT, B53, B54, B56) in solution

–> no introduction to confocal mode: nanobody not observable

08/14/2016

Sample preparation 7: GOPTS + anti-GFP nanobody + B53/B54

incubated with nanobody (1µl, 330µg/ml, 60 min)
rinsed and blocked with BSA(4%) (60 min)
rinsed and washed twice with PBS(1x) (10 min, 10 rpm)
rinsed with ddH2O
dried with compressed air
incubated with spores (10µl/spot, 120 min)
rinsed and washed twice with PBS(1x) (10 min, 10rpm)
rinsed with ddH20
dried with compressed air
incubation with Mowiol DABCO (5µl, overnight)

Spot No.	Sample
1	nanobody + WT (negative control)
2	nanobody + B53
3	nanobody + B54
4	nanobody + GFP (positive control)
5	nanobody (positive control)
6	GFP (positive control)

Washing test A: GOPTS + anti-GFP nanobody + B53

incubation with nanobody (1µl/mg, 300µg/ml, overnight, 4°C)

08/15/2016

Washing test A: GOPTS + anti-GFP nanobody + B53

rinsed and blocked with BSA(2%) (60 min, 10 rpm)
rinsed with PBS(1x), washed PBS twice (10 min, 10 rpm)
rinsed with ddH2O
dried with compressed air
incubation with B53 (10µl/spot, overnight)

08/16/2016

Washing test A: GOPTS + anti-GFP nanobody + B53

rinsed and washed (see table below)
rinsed with ddh2O
dried with compressed air
incubated with Mowiol DABCO (5µl, overnight)

Spot No.	Sample	Condition
1	GOPTS + nanobody + B53	washed 2x with PBS(1x) (10 min, 10 rpm)
2	GOPTS + nanobody + B53	washed 3x with PBS(1x) (10 min, 10 rpm)
3	GOPTS + nanobody + B53	washed 4x with PBS(1x) (10 min, 10 rpm)
4	GOPTS + nanobody + B53	washed 1x with PBS(1x) + 0,05% Tween20 (10 min, 10 rpm)
5	GOPTS + nanobody + B53	washed 1x with PBS(1x) + 0,1 % Tween20 (10 min, 10 rpm)

08/17/2016

Fluorescence microscopy 7 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + B53/B54

–> spores have not bound to nanobody

Fluorescence microscopy A (Nikon C2+ confocal microscope): Washing test: GOPTS + anti-GFP nanobody + B53

–> spores have not bound to BSA

08/21/2016

Sample preparation 8: GOPTS + anti-GFP nanobody + GFP

incubation with nanobody (see table below, 60 min)
rinsed and blocked with BSA(4%) (60 min)
rinsed and washed with PBS(1x) (5 min, 10 rpm)
rinsed with ddH2O
dried with compressed air
incubated with GFP (see table below, 1.35mg/ml, 20 min)
rinsed and washed with PBS(1x) (10 min, 10rpm)
rinsed with ddH20
dried with compressed air
incubation with Mowiol DABCO (5µl, overnight)

Spot No.	Sample
1	GOPTS + GFP (1µl) (positive control)
2	GOPTS + GFP (5 µl) (positive control)
3	GOPTS + nanobody (1µl, 330µg/ml) + GFP
4	GOPTS + nanobody (0.5µl, 10mg/ml) + GFP
5, 6	GOPTS + BSA + GFP (negative control)

08/22/2016

Fluorescence microscopy 8 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + GFP

–> GFP not bound, GOPTS plates too old

08/23/2016

Sample preparation 9: GOPTS + GFP

incubation with GFP (1.35mg/ml, volume/incubation: see table below)
rinsed and blocked with BSA(4%) (30 min)
rinsed and washed with PBS(1x) (10 min, 10 rpm)
rinsed with ddH2O
dried with compressed air
incubation with Mowiol DABCO (5µl, overnight)

Spot No.	Sample
1	GOPTS + GFP (1 µl, 10 min)
2	GOPTS + GFP (2 µl, 10 min)
3	GOPTS + GFP (5 µl, 10 min)
4	GOPTS + GFP (1 µl, 15 min)
5	GOPTS + GFP (2 µl, 15 min)
6	GOPTS + GFP (5 µl, 15 min)
7	GOPTS + GFP (1 µl, 20 min)
8	GOPTS + GFP (2 µl, 20 min)
9	GOPTS + GFP (5 µl, 20 min)
10	GOPTS (negative control)

Sample preparation 10: GOPTS + anti-GFP nanobody + B53

incubation with nanobody (1µl, 330µg/ml, 60 min)
rinsed and blocked with BSA(4%) (60 min)
rinsed and washed with PBS(1x) (10 min, 10 rpm)
rinsed with ddH2O
dried with compressed air
incubated with spores (10µl/spot, 30 min)
rinsed and washed with PBS(1x) (10min, 10rpm)
rinsed with ddH20
dried with compressed air
incubation with Mowiol DABCO (5µl, overnight)

Spot No.	Sample
1	GOPTS + nanobody + B53
2	GOPTS + nanobody + B53
3	GOPTS + GFP(positive control)
4	GOPTS + nanobody + GFP (positive control)
5	GOPTS + BSA + B53 (negative control)

Washing test B: GOPTS + anti-GFP nanobody + B53

incubation with nanobody (1µl, 330µg/ml, 60 min)
rinsed and blocked with BSA(4%) (60 min)
rinsed and washed with PBS(1x) (10 min, 10 rpm)
rinsed with ddH2O
dried with compressed air
incubated with spores (10µl/spot, 30 min)
rinsed and washed with PBS(1x) (10 min, 10rpm)
rinsed with ddH20
dried with compressed air
incubation with Mowiol DABCO (5µl, overnight)

Spot No.	Sample	Condition
1	GOPTS + nanobody + B53	rinsed with PBS(1x)
2	GOPTS + nanobody + B53	swivel with PBS(1x) (0.5 min)
3	GOPTS + nanobody + B53	washed with PBS(1x) (1 min, 10 rpm)
4	GOPTS + nanobody + B53	washed with PBS(1x) (2 min, 10 rpm)
5	GOPTS + nanobody + B53	washed with PBS(1x) (3 min, 10 rpm)
6	GOPTS + nanobody + B53	washed with PBS(1x) (5 min, 10 rpm)
7	GOPTS + BSA + B53(negative control)	washed with PBS(1x) (10 min, 10 rpm)
8	GOPTS + BSA + GFP(negative control)	washed with PBS(1x) (10 min, 10 rpm)
9	GOPTS + GFP (positive control)	washed with PBS(1x) (10 min, 10 rpm)
10	GOPTS + B53 (positive control)	washed with PBS(1x) (10 min, 10 rpm)

08/24/2016

Fluorescence microscopy 9 (Nikon C2+ confocal microscope): GOPTS + GFP

–> strong fluorescence signal at some of the edges: incubation too long/volume too little, GFP has dried and could not have been washed away properly

1) GOPTS + GFP (1 µl, 10 min)
2) GOPTS + GFP (2 µl, 10 min)
3) GOPTS + GFP (5 µl, 10 min)
4) GOPTS + GFP (1 µl, 15 min)
5) GOPTS + GFP (2 µl, 15 min)
6) GOPTS + GFP (5 µl, 15 min)
7) GOPTS + GFP (1 µl, 20 min)
8) GOPTS + GFP (2 µl, 20 min)
9) GOPTS + GFP (5 µl, 20 min)

Fluorescence microscopy 10 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + B53

–> no spores appear to have bound to nanobody

Fluorescence microscopy B (Nikon C2+ confocal microscope): Washing test: GOPTS + anti-GFP nanobody + B53

–> no spores after washing, no binding between nanobody and B53?

08/25/2016

Blocking test C: GOPTS + BSA/milk powder + GFP

blocking with BSA(4%)/milk powder(5%) (120 min)
rinsed and washed with PBS(1x) (10 min, 10 rpm)
incubation with GFP (1µl/spot, 1.35mg/ml, 20 min)
rinsed and washed with PBS(1x) (10 min, 10 rpm)
rinsed with ddH20
dried with compressed air
incubation with Mowiol DABCO (5µl, overnight)

Spot No.	Sample
1	GOPTS + GFP (positive control)
2	GOPTS + milk powder(5%) + GFP
3	GOPTS + BSA(4%) + GFP

Contamination Test D: BSA, PBS Mowiol DABCO

–> no contamination/pollution detected

Sample preparation 11: GOPTS + mCherry

incubated with mCherry (1µl, 2mg/ml, incubation: see table below)
rinsed and blocked with BSA(4%) (30 min)
rinsed and washed with PBS(1x) (10 min, 10 rpm)
rinsed with ddH2O
dried with compressed air
incubated with Mowiol DABCO (5µl, overnight)

Spot No.	Sample
1	GOPTS + mCherry (5 min)
2	GOPTS + mCherry (10 min)
3	GOPTS + mCherry (15 min)
4	GOPTS (negative control)
5	mCherry + Mowiol DABCO (positive control)

Sample preparation 12: GOPTS + anti-GFP nanobody + GFP

incubated with nanobody (5µl, 330µg/ml, 120 min)
rinsed and blocked with milk powder(5%) (60 min)
rinsed and washed with PBS(1x) (10 min, 10 rpm)
rinsed with ddH2O
dried with compressed air
incubated with GFP (2µl, 1.35mg/ml, 20 min)
rinsed and washed with PBS(1x) (10 min, 10rpm)
rinsed with ddH20
dried with compressed air
incubated with Mowiol DABCO (5µl, overnight)

Spot No.	Sample
1	GOPTS + nanobody + GFP
2	GOPTS (negative control)
3	GOPTS + GFP (positive control)
4	GOPTS + nanobody (negative control)
5	GOPTS + BSA + GFP (negative control)

08/27/2016

Fluorescence microscopy C (Nikon C2+ confocal microscope): Blocking test: GOPTS + BSA/milkpowder + GFP

–> No GFP signal after blocking with BSA and milk powder

Fluorescence microscopy 11 (Nikon C2+ confocal microscope): GOPTS + mCherry

–> no mCherry-signal, too weak (quantum yield of mCherry about 1/3 of that of GFP)

Fluorescence microscopy 12 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + GFP

–> GFP has not bound to nanobody

08/30/2016

Sample preparation 13: GOPTS + nanobody + GFP

incubated with nanobody (5µl, 330µg/ml, 60 min)
rinsed and blocked with milk powder(5%) (30 min)
rinsed and washed with PBS(1x) (10 min, 10 rpm)
rinsed with ddH2O
dried with compressed air
incubated with GFP (2µl, 1.35mg/ml, 20 min)
rinsed and washed with PBS(1x) (10 min, 10rpm)
rinsed with ddH20
dried with compressed air
incubated with Mowiol DABCO (5µl, overnight)

Spot No.	Sample
1,2	GOPTS + nanobody(Alexa647) + GFP
3,4	GOPTS + nanobody + GFP
5	GOPTS + GFP (positive control)
6	GOPTS + nanobody (negative control)
7	GOPTS + BSA (negative control)

09/01/2016

Fluorescence microscopy 13 (Nikon C2+ confocal microscope): GOPTS + nanobody + GFP

–> no GFP signal detectable

09/06/2016

Sample preparation 14: GOPTS + GFP

incubated with GFP (10µl, 1.35mg/ml, 30 min)
rinsed and blocked with milk powder(5%) (30 min)
rinsed and washed with PBS(1x) (10 min, 10 rpm)
rinsed with ddH2O
dried with compressed air
incubated with Mowiol DABCO (5µl, overnight)

Spot No.	Sample
1,2	GOPTS + mCherry
3	GOPTS + BSA + mCherry (negative control)
4	GOPTS + BSA (negative control)
5	mCherry (positive control)

Mowiol was produced due to this protocol: https://www.carlroth.com/downloads/ba/en/0/BA_0713_EN.pdf

09/07/2016

Fluorescence microscopy 14 (Nikon C2+ confocal microscope): GOPTS + GFP

–> spots with evenly distributed GFP

1) GOPTS + GFP
2) GOPTS + nanobody + GFP

09/14/2016

Sample preparation 16: GOPTS + B53

incubated with B53 (5µl, 60 min)
rinsed and blocked with milk powder(4%) (30 min)
rinsed and washed with PBS(1x) (10 min, 10 rpm)
rinsed with ddH2O
dried with compressed air
incubated with Mowiol DABCO (5µl, overnight)

Spot No.	Sample
1,2,3	GOPTS + B53
4	GOPTS + WT (negative control)
5	WT (negative control)
6	B53 (positive control)
7	GOPTS + GFP

09/15/2016

Sample preparation 17: GOPTS + anti-GFP nanobody(Alexa647) + GFP

incubated with nanobody (5µl, 330µg/ml, 60 min)
rinsed and blocked with milk powder(5%) (30 min)
rinsed and washed with PBS(1x) (10 min, 10 rpm)
rinsed with ddH2O
dried with compressed air
incubated with GFP (2µl, 1.35mg/ml, 20 min)
rinsed and washed with PBS(1x) (10 min, 10rpm)
rinsed with ddH20
dried with compressed air
incubated with Mowiol DABCO (5µl, overnight)

Spot No.	Sample
1-4	GOPTS + nanobody(Alexa647) + GFP
5	GOPTS + milk powder + GFP (negative control)
6	GOPTS + milk powder + NB (negative control)
7	GOPTS + GFP
8	GOPTS (negative control)

Fluorescence microscopy 16 (Nikon C2+ confocal microscope): GOPTS + B53

–> no spores detectable

09/16/2016

Fluorescence microscopy 17 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + GFP

–> GFP signal: GFP has been bound to nanobody

1-4) GOPTS + nanobody(Alexa647) + GFP
5) GOPTS + milk powder + GFP (negative control)
6) GOPTS + milk powder + NB (negative control)
7) GOPTS + GFP (positive control)
8) GOPTS (negative control)

09/17/2016

Sample preparation 18: GOPTS + anti-GFP nanobody + GFP (spotting-mask)

incubation with anti-GFP nanobody ( 2µl/spot, 330µg/ml, 10 min, 4°C)
rinsing and blocking with milk powder(5%) (15 min, 10 rpm)
rinsing with PBS(1x)
incubated with GFP (2µl, 1.35mg/ml, 30 min, 4°C)
rinsed and washed with PBS(1x) (10 min, 10 rpm)
rinsed with ddH2O
(removed mask)
rinsed with ddH2O
dried with compressed air
incubated with Mowiol DABCO (overnight)

09/18/2016

Fluorescence microscopy 18 (Nikon C2+ confocal microscope): GOPTS + antiGFP-nanobody + GFP (spotting-mask)

–> no GFP signal detectable

Sample preparation 19: GOPTS + anti-GFP nanobody + GFP (spotting-mask)

incubation with anti-GFP nanobody (2µl/spot, 330µg/ml, 15 min, 4°C)
rinsing with PBS(1x)
(removed mask)
rinsing and blocking with milk powder(5%) (15 min, 2 rpm)
(reattached mask)
incubated with GFP (2µl, 1.35mg/ml, 30 min, 4°C)
(removed mask)
rinsed and washed with PBS(1x) (10 min, 10 rpm)
rinsed with ddH2O
dried with compressed air
incubated with Mowiol DABCO (overnight)

09/19/2016

Fluorescence microscopy 19 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + GFP (spotting-mask)

–> slight GFP-signal on nanobody

Sample preparation 20: anti-GFP nanobody + B53 (in solution)

incubation of anti-GFP nanobody(Alexa647) (2µl, 165µg/ml, 15 min, 4°C) + B53
incubated on GOPTS-slide (2µl, 15 min)
rinsed and blocked with milk powder(5%) (30 min, 10 rpm)
rinsed and washed with PBS(1x) (2 x 1 min, 10 rpm)

–> problem: no washing away of unbound nanobody

Spot No.	Sample
1,2	B53 + nanobody(Alexa647) (incubated in solution) on GOPTS
3,4	WT + nanobody(Alexa647) (incubated in solution) on GOPTS (negative control)
5,6	B53 + nanobody(Alexa647) (incubated in solution) dried on milk powder
7,8	WT + nanobody(Alexa647) (incubated in solution) dried on milk powder (negative control)
9,10	GOPTS + nanobody(Alexa647) + GFP (positive control)
11,12	GOPTS + nanobody(Alexa647) + B53
13,14	GOPTS + nanobody(Alexa647) + WT (negative control)
15	GOPTS + milk powder + nanobody(Alexa647) (negative control)
16	GOPTS + milk powder + B53 (negative control)
17	GOPTS + milk powder + WT (negative control)
18	GOPTS + GFP (positive control)
19	GOPTS + nanobody(Alexa647) (negative control)
20	B53 dried (positive control)

09/20/2016

Sample preparation 21: anti-GFP nanobody + B53 (in solution)

incubation with anti-GFP nanobody(Alexa647) (2µl, 165µg/ml, 30 min, 4°C) + B53
centrifuged (2 min, 13400 rpm)
washed with PBS(1x)
repeated steps 2 and 3
resuspended in PBS(1x) (30µl)
incubated on GOPTS-slide (2µl, 15 min)
rinsed and blocked with milkpowder(5%) (30 min, 10 rpm)
rinsed and washed with PBS(1x) (2 x 1 min, 10 rpm)

Spot No.	Sample
1	B53 + nanobody(Alexa647) (incubated in solution) on GOPTS
2	WT + nanobody(Alexa647) (incubated in solution) on GOPTS
3,4	B53 + nanobody(Alexa647) dried on milkpowder
5,6	WT + nanobody(Alexa647) dried on milkpowder

09/29/2016

Sample preparation 22: GOPTS + GFP

incubation with GFP (1 µl, 1.35mg/ml, 30 min)
rinsed and blocked with milk powder(5%) (30 min)
rinsed and washed with PBS(1x) (10 min, 10 rpm)
rinsed with ddH2O
dried with compressed air
incubation with Mowiol DABCO (5µl, overnight)

Spot No.	Sample
1-5	GOPTS + GFP
6	GOPTS

09/30/2016

–> Construct spores are ready for analysis

Strain No.	Strain
3	150: aGFPnano_HA_aHelix_cgeA
6	151: aGFPnano_HA_aHelix_cotG
7	159: aGFPnano_HA_G4S_CotZ
9	157: aGFPnano_HA_aHelix_CotZ

Fluorescence microscopy 22 (Nikon C2+ confocal microscope): GOPTS + GFP

–> spots with evenly distributed GFP

incubation with GFP (1 µl, 1.35mg/ml, 30 min)
spores centrifuged (2 min, 13,400 rpm)
washed with PBS(1x)
step 1 repeated
resuspended in PBS(1x)
incubation on GOPTS-slide(GFP) with spores (4µl, 400,000/µl, incubation time: see table below, 4°C)
rinsed and blocked with milk powder(5%) (30 min, 10 rpm)
rinsed with PBS(1x)
rinsed with ddH2O
dried with compressed air
incubation with Mowiol DABCO (overnight)

Spot No.	Sample	Incubation time (min)
1	GOPTS + GFP + strain 3	5
2	GOPTS + GFP + strain 3	10
3	GOPTS + GFP + strain 3	15
4	GOPTS + GFP + strain 6	5
5	GOPTS + GFP + strain 6	10
6	GOPTS + GFP + strain 6	15
7	GOPTS + GFP + strain 7	5
8	GOPTS + GFP + strain 7	10
9	GOPTS + GFP + strain 7	15
10	GOPTS + GFP + strain 9	5
11	GOPTS + GFP + strain 9	10
12	GOPTS + GFP + strain 9	15
13, 14	GOPTS + GFP (positive control)	10
15	GOPTS + GFP + WT (negative control)	5
16	GOPTS + GFP + WT (negative control)	10
17	GOPTS + GFP + WT (negative control)	15
18	GOPTS + strain 3 (positive control)	10
19	GOPTS + strain 6 (positive control)	10
20	GOPTS + strain 7 (positive control)	10
21	GOPTS + strain 9 (positive control)	10
22	GOPTS + WT (negative control)	10

10/01/2016

Sample preparation 23: GOPTS + GFP + spores

–> no spores detectable

Sample preparation 24: GOPTS + GFP + spores (in EtOH resuspended)

incubation of GFP (4µl, 1,35g/ml) + XXX (4µl, 400,000/µl) (10 min, 4°C)
solution centrifuged (4 min, 13,400 rpm)
washed with PBS(1x)
step 2 repeated
resuspended in EtOH (250µl)
incubation on GOPTS-slide (4µl, 30 min, 4°C)
rinsed and blocked with milk powder(5%) (30 min, 10 rpm)
rinsed with PBS(1x)
rinsed with ddH2O
dried with compressed air
incubation with Mowiol DABCO (overnight)

Spot No.	Sample
1-3	GOPTS + GFP + strain 6
4-6	GOPTS + GFP + WT
7-9	GOPTS + GFP
10-12	GOPTS + strain 6
13-15	GOPTS + WT

10/02/2016

Fluorescence microscopy 24 (Nikon C2+ confocal microscope): GOPTS + GFP + spores (strains 3,6,7,9,wt) (in EtOH resuspended)

–> EtOH denaturated GFP: very weak signal. No spores detectable

Sample preparation 25: GFP + spores (strains 3,6,7,9) (in solution)

incubation of GFP (4µl, 1,35g/ml) + spores(strain 3,6,7,9) (4µl, 400,000/µl) (incubation time: see table below, 4°C)
centrifuged (2 min, 13400 rpm)
washed with PBS(1x)
repeated steps 2 and 3
resuspended in PBS(1x) (20µl)
incubated on GOPTS-slide (4µl, 10 min)
rinsed and blocked with milk powder(5%) (30 min, 10 rpm)
rinsed and washed with PBS(1x) (10 min, 10 rpm)
rinsed with ddH2O
dried with compressed air
incubrnight)ation with Mowiol DABCO

Spot No.	Sample
1, 2	GFP + strain 3
3, 4	GFP + strain 6
5, 6	GFP + strain 7
7, 8	GFP + strain 9
9, 10	GFP + WT (negative control)
11, 12	GFP + PBS (positive control)
13, 14	GFP (positive control)
15, 16	WT (negative control)
17	strain 3 (negative control)
18	strain 6 (negative control)
19	strain 7 (negative control)
20	strain 9 (negative control)

10/03/2016

Sample preparation 26: Incubation test: GFP + spores (in solution)

incubation of GFP (4µl, 1,35g/ml) + spores (4µl, 400,000/µl) (incubation time: see tables below, 4°C)
centrifuged (2 min, 13400 rpm)
washed with PBS(1x)
repeated steps 2 and 3)
resuspended in PBS(1x) (20µl))
incubated on GOPTS-slide (4µl, 10 min)
rinsed and blocked with milk powder(5%) (30 min, 10 rpm)
rinsed and washed with PBS(1x) (10 min, 10 rpm)
rinsed with ddH2O
dried with compressed air
incubation with Mowiol DABCO (overnight)

Slide 1:

Spot No.	Sample	Incubation time
1, 2	GFP + strain 3	30 min
3, 4	GFP + strain 3(pellet washed with PBS(1x))	30 min
5, 6	GFP + strain 6	30 min
7, 8	GFP + strain 6(pellet washed with PBS(1x))	30 min
9, 10	GFP + strain 7	30 min
11, 12	GFP + strain 7(pellet washed with PBS(1x))	30 min
13, 14	GFP + strain 9	30 min
15, 16	GFP + strain 9 (pellet washed withn PBS(1x))	30 min
17, 18	GFP + WT (negative control)
19, 20	GFP + WT (negative control) (pellet washed with PBS(1x))
21, 22	GFP (positive control)
23, 24	WT (negative control)
25	strain 3 (negative control)
26	strain 6 (negative control)
27	strain 7 (negative control)
28	strain 9 (negative control)

Slide 2:

Spot No.	Sample	Incubation time
1, 2	GFP + strain 3	60 min
3, 4	GFP + strain 3(pellet washed with PBS(1x))	60 min
5, 6	GFP + strain 6	60 min
7, 8	GFP + strain 6(pellet washed with PBS(1x))	60 min
9, 10	GFP + strain 7	60 min
11, 12	GFP + strain 7(pellet washed with PBS(1x))	60 min
13, 14	GFP + strain 9	60 min
15, 16	GFP + strain 9 (pellet washed withn PBS(1x))	60 min
17, 18	GFP + WT (negative control)
19, 20	GFP + WT (negative control) (pellet washed with PBS(1x))
21, 22	GFP (positive control)
23, 24	WT (negative control)
25	strain 3 (negative control)
26	strain 6 (negative control)
27	strain 7 (negative control)
28	strain 9 (negative control)

Slide 3:

Spot No.	Sample	Incubation time
1, 2	GFP + strain 3	120 min
3, 4	GFP + strain 3(pellet washed with PBS(1x))	120 min
5, 6	GFP + strain 6	120 min
7, 8	GFP + strain 6(pellet washed with PBS(1x))	120 min
9, 10	GFP + strain 7	120 min
11, 12	GFP + strain 7(pellet washed with PBS(1x))	120 min
13, 14	GFP + strain 9	120 min
15, 16	GFP + strain 9 (pellet washed withn PBS(1x))	120 min
17, 18	GFP + WT (negative control)
19, 20	GFP + WT (negative control) (pellet washed with PBS(1x))
21, 22	GFP (positive control)
23, 24	WT (negative control)
25	strain 3 (negative control)
26	strain 6 (negative control)
27	strain 7 (negative control)
28	strain 9 (negative control)

Sample preparation 28: GFP-volume test B: GFP + strain 7 (in solution, washed)

incubation of GFP (volume: see table below, 1,35g/ml) + strain 7 (4µl, 400,000/µl) (90 min, 4°C)
centrifuged (2 min, 13400 rpm)
washed with PBS(1x)
repeated steps 2 and 3)
resuspended in PBS(1x) (20µl)
incubated on GOPTS-slide (2µl, 90 min)
rinsed and blocked with milk powder(5%) (30 min, 10 rpm)
rinsed and washed with PBS(1x) (10 min, 10 rpm)
rinsed with ddH2O
dried with compressed air
incubation with Mowiol DABCO (overnight)

Spot No.	Sample	Volume (GFP)
1, 2	GFP + strain 7	0,5µl
3, 4	GFP + strain 7	1,0µl
5, 6	GFP + strain 7	1,5µl
7, 8	GFP + strain 7	2,0µl
9, 10	GFP + strain 7	2,5µl
11, 12	GFP (positive control)	4,0µl
13, 14	strain 7 (negative control)
15, 16	GFP + WT (negative control)	0,5µl
17, 18	GFP + WT (negative control)	1,0µl
19, 20	GFP + WT (negative control)	1,5µl
21, 22	GFP + WT (negative control)	2,0µl
23, 24	GFP + WT (negative control)	2,5µl
25, 26	WT (negative control)

Sample preparation 29: Spore-amount test: GFP + strain 7 (in solution)

incubation of GFP (0,5µl, 1,35g/ml) + strain 7 (amount: see table below, 400,000/µl) (90 min, 4°C)
centrifuged (2 min, 13400 rpm)
washed with PBS(1x)
repeated steps 2 and 3)
resuspended in PBS(1x) (20µl)
incubated on GOPTS-slide (2µl, 90 min)
rinsed and blocked with milk powder(5%) (30 min, 10 rpm)
rinsed and washed with PBS(1x) (10 min, 10 rpm)
rinsed with ddH2O
dried with compressed air
incubation with Mowiol DABCO (overnight)

Spot No.	Sample	Amount (spores)
1, 2	strain 7 (negative control)	25,000,000
3, 4	strain 7 (negative control)	15,000,000
5, 6	strain 7 (negative control)	10,000,000
7, 8	GFP + strain 7	25,000,000
9, 10	GFP + strain 7	15,000,000
11, 12	GFP + strain 7	10,000,000
13, 14	GFP (positive control)
15, 16	WT (negative control)	25,000,000
17, 18	WT (negative control)	15,000,000
19, 20	WT(negative control)	10,000,000
21, 22	GFP + WT (negative control)	25,000,000
23, 24	GFP + WT (negative control)	15,000,000
25, 26	GFP + WT (negative control)	10,000,000

10/04/2016

Fluorescence microscopy 24 (Nikon C2+ confocal microscope): GOPTS + GFP + spores (in EtOH resuspended)

–> spore concentration too high, too many layers of spores

Fluorescence microscopy 25 (Nikon C2+ confocal microscope): GFP + spores (in solution)

–> spore concentration too high, too many layers of spores

Fluorescence microscopy 26 (Nikon C2+ confocal microscope): Incubation test: GFP + spores (in solution)

–> spore concentration too high, too many layers of spores

Slide 1:

1.1) strain7 + GFP (30 min) (white field) 1.2) strain7 + GFP (30 min) (GFP-channel) 2.1) strain9 + GFP (30 min) (white field) 2.2) strain9 + GFP (30 min) (GFP-channel)

Slide 2:

1.1) strain 7 + GFP (90 min)(white field) 1.2) strain 7 + GFP (90 min)(GFP-channel) 2.1) strain WT + GFP (90 min)(white field) 2.2) strain WT + GFP (90 min)(GFP-channel)

Fluorescence microscopy 27 (Nikon C2+ confocal microscope): GFP-volume test A: GFP + strain 7 (in solution)

–> background fluorescence was too high

Fluorescence microscopy 28 (Nikon C2+ confocal microscope): GFP-volume test B: GFP + strain 7 (in solution)

–> background fluorescence was too high

Fluorescence microscopy 29 (Nikon C2+ confocal microscope): GFP + strain 7 (in solution)

–> spores and WT show similar levels of fluorescence, probably autofluorescence

1) strain 7 + GFP (white field) 2) strain 7 + GFP (GFP-channel) 3) WT + GFP (white field) 4) WT + GFP (GFP-channel)

Sample preparation 30: Wash-test A: GFP + spores (strains 3,6,7,9) (in solution)

incubation of GFP (1µl, 1,35g/ml) + spores (strain 3,6,7,9) (25,000.000 spores) (90 min, 4°C)
solution centrifuged (2 min, 13,400 rpm)
washed with PBS(1x)
step 2 and 3 repeated (2x/4x)
resuspended in EtOH (250µl)
incubation on GOPTS-slide (4µl, 30 min, 4°C)
rinsed and blocked with milk powder(5%) (30 min, 10 rpm)
rinsed with PBS(1x)
rinsed with ddH2O
dried with compressed air
incubation with Mowiol DABCO (overnight)

Spot No.	Sample	Washing of GFP-spore solution
1, 2	GFP + strain 3	2x
3, 4	GFP + strain 6	2x
5, 6	GFP + strain 7	2x
7, 8	GFP + strain 9	2x
9, 10	GFP + WT (negative control)	2x
11, 12	GFP + strain 3	4x
13, 14	GFP + strain 6	4x
15, 16	GFP + strain 7	4x
17, 18	GFP + strain 9	4x
19, 20	GFP + WT (negative control)	4x
21, 22	GFP (positive control)
23	WT (negative control)
24	strain 3 (negative control)
25	strain 6 (negative control)
26	strain 7 (negative control)
27	strain 9 (negative control)

Sample preparation 31: Wash-test B: GFP + spores (in solution)

incubation of GFP (1µl, 1,35g/ml) + spores (25,000.000 spores) (90 min, 4°C)
solution centrifuged (2 min, 13,400 rpm)
washed with TBST(0,005%)
steps 2 and 3 repeated (2x/4x)
resuspended in EtOH (250µl)
incubation on GOPTS-slide (4µl, 30 min, 4°C)
rinsed and blocked with milk powder(5%) (30 min, 10 rpm)
rinsed with PBS(1x)
rinsed with ddH2O
dried with compressed air
incubation with Mowiol DABCO (overnight)

Spot No.	Sample	Washing of GFP-spore solution
1, 2	GFP + strain 3	2x
3, 4	GFP + strain 6	2x
5, 6	GFP + strain 7	2x
7, 8	GFP + strain 9	2x
9, 10	GFP + WT (negative control)	2x
11, 12	GFP + strain 3	4x
13, 14	GFP + strain 6	4x
15, 16	GFP + strain 7	4x
17, 18	GFP + strain 9	4x
19, 20	GFP + WT (negative control)	4x
21, 22	GFP (positive control)
23	WT (negative control)
24	strain 3 (negative control)
25	strain 6 (negative control)
26	strain 7 (negative control)
27	strain 9 (negative control)

10/05/2016

Fluorescence microscopy 30 (Nikon C2+ confocal microscope): Wash-test A: GFP + spores (strains 3,6,7,9) (in solution)

–> spore concentration too high, background fluorescence too high

Fluorescence microscopy 31 (Nikon C2+ confocal microscope): Wash-test B: GFP + spores (strains 3,6,7,9) (in solution)

–> indicates that strain 3 binds GFP

1.1) GFP + strain 3 (white field)
1.2) GFP + strain 3 (GFP-channel)
2.1) GFP + WT (negative control) (white field)
2.2) GFP + WT (negative control) (GFP-channel)
3) WT (negative control)

10/06/2016

Sample preparation 32: Wash-test C: GFP + spores (in solution)

incubation of GFP (1µl, 1,35g/ml) + spores (25,000.000 spores) (90 min, 4°C)
solution centrifuged (2 min, 13,400 rpm)
washed with TBST(0,005%)
steps 2 and 3 repeated (1x/3x/5x)
resuspended in EtOH (250µl)
incubation on GOPTS-slide (4µl, 30 min, 4°C)
rinsed and blocked with milk powder(5%) (30 min, 10 rpm)
rinsed with PBS(1x)
rinsed with ddH2O
dried with compressed air
incubation with Mowiol DABCO (overnight)

Spot No.	Sample	Washing of GFP-spore solution (step 3)
1, 2	GFP + strain 3	1x
3, 4	GFP + strain 7	1x
5, 6	GFP + WT	1x
7, 8	GFP + strain 3	3x
9, 10	GFP + strain 7	3x
11, 12	GFP + WT	3x
13, 14	GFP + strain 3	5x
15, 16	GFP + strain 7	5x
17, 18	GFP + WT	5x
19, 20	WT (negative control)
21, 22	GFP (positive control)

10/07/2016

Fluorescence microscopy 32 (Nikon C2+ confocal microscope): Wash-test B: GFP + spores (in solution)

–> spores and WT show similar levels of fluorescence, probably autofluorescence

Sample preparation 33: Blocking-test A: spores milk powder + GFP (in solution)

solution centrifuged (2 min, 13,400 rpm)
spores washed twice with PBS(1x) (2x 100µl)
spores resuspended and incubated in milk powder(5%) (100µl, incubation time: see table below)
incubation with GFP (5µl, 1,35g/ml, 90 min)
washed with PBS(1x) (3x 100µl)
resuspended in PBS(1x) (20 µl)
incubation on GOPTS-slide (4µl, 30 min, 4°C)
rinsed and blocked with milk powder(5%) (30 min, 10 rpm)
rinsed with PBS(1x)
rinsed with ddH2O
dried with compressed air
incubation with Mowiol DABCO (overnight)

Spot No.	Sample	Incubation time
1, 2	strain 3 + milk powder + GFP	30 min
3, 4	strain 3 + milk powder + GFP	60 min
5, 6	strain 7 + milk powder + GFP	30 min
7, 8	strain 7 + milk powder + GFP	60 min
9, 10	WT + milk powder + GFP (negative control)	30 min
11, 12	WT + milk powder + GFP(negative control)	60 min
13, 14	WT + milk powder (negative control)	30 min
15, 16	WT + milk powder (negative control)	60 min
17, 18	strain 3 + milk powder (negative control)	30 min
19, 20	strain 3 + milk powder (negative control)	60 min
21, 22	strain 7 + milk powder (negative control)	30 min
23, 24	strain 7 + milk powder (negative control)	60 min
25, 26	GFP (positive control)	30 min

10/08/2016

Fluorescence microscopy 33 (Nikon C2+ confocal microscope): Blocking-test A: spores + milk powder + GFP (in solution)

–> spores and WT show similar levels of fluorescence

10/11/2016

Sample preparation 34: Blocking-test B: spores + BSA + GFP (in solution)

solution centrifuged (2 min, 13,400 rpm)
spores (5,000,000) washed with PBS(1x) (3x 100µl)
spores resuspended and incubated in BSA(5%) (100µl, 60 min)
incubation with GFP (50µl, 100µg/ml, 60 min)
washed with TBST(0.05%) (5x 100µl)
resuspended in PBS(1x) (50 µl)
incubation with Mowiol DABCO (overnight)

Spot No.	Sample
1, 2	strain 3 + BSA + GFP
3, 4	strain 7 + BSA + GFP
5, 6	WT + BSA + GFP (negative control)
7, 8	WT + BSA (negative control)
9, 10	strain 3 + milk powder (negative control)
11, 12	strain 7 + milk powder (negative control)
13, 14	GFP (positive control)

10/12/2016

Fluorescence microscopy 34 (Nikon C2+ confocal microscope): Blocking-test B: spores + BSA + GFP (in solution)

–> too little spores

lab book

@@ Line 1: / Line 1: @@
 {{Freiburg/Styles}}
+{{Freiburg/Menubar}}
 {{Freiburg/Away}}
-{{Freiburg/Menubar}}
 {{Freiburg/Main}}
+{{Freiburg/Top}}
 <html>
-<style>
- .image img:hover {
-        cursor: pointer;
-        -webkit-transform: scale(1.1);
-        /* Safari and Chrome */
-        -moz-transform: scale(1.1);
-        /* Firefox */
-        transition-duration: .2s;
-        transition-timing-function: linear;
-    }
-</style>
-<div class="" >
-<h5   style="text-align: center"> Lab Journals </h5>
-<center>
-<div class="image">
-             <a href="https://2016.igem.org/Team:Freiburg/NotebookCloning"> <img class="imga" src="https://static.igem.org/mediawiki/2016/1/14/T--Freiburg--CloningLab.png" alt="Smiley face"style="width:200px"> </a>
-            <a href="https://2016.igem.org/Team:Freiburg/NotebookSpores"> <img class="imgb" src="https://static.igem.org/mediawiki/2016/9/93/T--Freiburg--SporesLab.png" alt="Smiley face" style="width:200px"> </a>
-            <a href="https://2016.igem.org/Team:Freiburg/NotebookProteins"> <img class="imgc" src="https://static.igem.org/mediawiki/2016/2/23/T--Freiburg--ProteinsLab.png" alt="Smiley face" style="width:200px"> </a>
-            <a href="https://2016.igem.org/Team:Freiburg/NotebookTargeting"> <img class="imgd" src="https://static.igem.org/mediawiki/2016/f/f4/T--Freiburg--TargetingLab.png" alt="Smiley face" style="width:200px"> </a>
-            <a href="https://2016.igem.org/Team:Freiburg/NotebookDelivery"> <img class="imge" src="https://static.igem.org/mediawiki/2016/4/4b/T--Freiburg--DeliveryLab.png" alt="Smiley face" style="width:200px"> </a>
-</div>
-</div>
-</center>
-<div class="para_center_20">
-<h1 class="sectionedit1"><a name="group_4" id="group_4">Group 4</a></h1>
-<div class="level1">
-<p>
-<strong>Members:Kevin, Christian, Vivi, Nathalie</strong><br/>
-</p>
-</div>
-<h4><a name="section07042016" id="section07042016">07/04/2016</a></h4>
-<div class="level4">
-<p>
-<strong>Preparation of overnight cultures of 5 different <em>Bacillus subtilis</em> strains with GFP on CotZ-fusionprotein</strong>
-</p>
-<div class="table sectionedit2"><table class="inline">
-	<tr class="row0">
-		<td class="col0">Nr.</td><td class="col1">Strain</td>
-	</tr>
-	<tr class="row1">
-		<td class="col0">1.</td><td class="col1">B52:B. Subtilis w168 amyE::PcgeA-CotZre-gfp-B0014(CM5)</td>
-	</tr>
-	<tr class="row2">
-		<td class="col0">2.</td><td class="col1">B53:B. Subtilis w168 amyE::PcotYZ-CotZre-gfp-B0014(CM5)</td>
-	</tr>
-	<tr class="row3">
-		<td class="col0">3.</td><td class="col1">B54:B. Subtilis w168 amyE::PcotYZ-CotZin-gfp-B0014(CM5)</td>
-	</tr>
-	<tr class="row4">
-		<td class="col0">4.</td><td class="col1">B55:B. Subtilis w168 amyE::PcotV-CotZre-gfp-B0014(CM5)</td>
-	</tr>
-	<tr class="row5">
-		<td class="col0">5.</td><td class="col1">B56:B. Subtilis w168 amyE::PCotv-CotZin-gfp-B0014(CM5)</td>
-	</tr>
-</table></div>
-<!-- EDIT2 TABLE [206-520] -->
-<p>
-<strong>Medium:</strong>
-Preparation of LB-medium with <em>Chloramphenicol</em>: 50µl(1000X) in 50ml LB-Medium<br/>
-</p>
-<p>
-<strong>Inoculation:</strong>
-reaction tubes with 5 ml LB-<em>Chloramphenicol</em>-medium have been used. Each has been inoculated with a little piece of filter paper with spores. Cultures have been incubated at 200rpm and 37°C, 16:10Pm to 8am.
-No growth has been observed !
-</p>
-<p>
-Further spore production was performed by group 2.
-</p>
-</div>
-<h4><a name="section07052016" id="section07052016">07/05/2016</a></h4>
-<div class="level4">
-<p>
-<strong>Preparation of GOPTS coated glass slides</strong>
-</p>
-<p>
-Surface activation of glass slides by plasma generator, coated with GOPTS ((3-glycidyloxypropyl)trimethoxysilane), washed with acetone and then stored for up to two weeeks.
-</p>
-<ol>
-<li class="level1"><div class="li"> labeled with a diamond pen</div>
-</li>
-<li class="level1"><div class="li"> washed with EtOH then ddH2O</div>
-</li>
-<li class="level1"><div class="li"> dried slides with compressed air</div>
-</li>
-<li class="level1"><div class="li"> activation of surface with plasma generator</div>
-</li>
-<li class="level1"><div class="li"> incubation with GOPTS (overnight)</div>
-</li>
-</ol>
-</div>
-<h4><a name="section07102016" id="section07102016">07/10/2016</a></h4>
-<div class="level4">
-<p>
-<strong>Buffer for anti-GFP-nanobody (10 ml)</strong>
-</p>
-<ul>
-<li class="level1"><div class="li"> 61mg Tris </div>
-</li>
-<li class="level1"><div class="li"> 87mg NaCl </div>
-</li>
-<li class="level1"><div class="li"> 1ml Glycerin </div>
-</li>
-<li class="level1"><div class="li"> 2,5 ml Imidazole(1M)</div>
-</li>
-<li class="level1"><div class="li"> fill up with ddH2O to 10 ml</div>
-</li>
-</ul>
-</div>
-<h4><a name="section07122016" id="section07122016">07/12/2016</a></h4>
-<div class="level4">
-<p>
-<strong>Sample preparation 1: GOPTS + anti-GFP nanobody + B52-B56</strong>
-</p>
-<ol>
-<li class="level1"><div class="li"> Glass slides were spotted with 0,5 µl nanobody (300µg/ml) (Spot location marked with permanent marker.)</div>
-</li>
-<li class="level1"><div class="li"> Slides were incubated over night in the fridge at 4°C in darkness</div>
-</li>
-<li class="level1"><div class="li"> 150µl BSA(2x) on each spot, 30 min incubation</div>
-</li>
-<li class="level1"><div class="li"> washed with PBS(1x)</div>
-</li>
-<li class="level1"><div class="li"> washed with 20mM imidazole in PBS(1x)</div>
-</li>
-<li class="level1"><div class="li"> washed with PBS(1x)</div>
-</li>
-<li class="level1"><div class="li"> washed with ddH2o</div>
-</li>
-<li class="level1"><div class="li"> dried with compressed air </div>
-</li>
-<li class="level1"><div class="li"> 100µl of spore solution on each slide</div>
-</li>
-</ol>
-</div>
-<h4><a name="section07132016" id="section07132016">07/13/2016</a></h4>
-<div class="level4">
-<p>
-<strong>Sample preparation 1: GOPTS + anti-GFP nanobody + B52-B56</strong>
-</p>
-<ol>
-<li class="level1"><div class="li"> blocked with BSA(2%) (150µl/spot, 30 min) </div>
-</li>
-<li class="level1"><div class="li"> washed with PBS(1x)</div>
-</li>
-<li class="level1"><div class="li"> washed with imidazole (20mM) in PBS(1x)</div>
-</li>
-<li class="level1"><div class="li"> washed with PBS(1x)</div>
-</li>
-<li class="level1"><div class="li"> washed with ddH2O</div>
-</li>
-<li class="level1"><div class="li"> dried with compressed air </div>
-</li>
-<li class="level1"><div class="li"> incubation with B52-B56 (100µl/spot, 120 min) </div>
-</li>
-<li class="level1"><div class="li"> incubated with Mowiol DABCO(1,4-Diazabicyclo[2.2.2]octane) (5µl/spot, overnight), with second glass slide on top </div>
-</li>
-</ol>
-<div class="table sectionedit3"><table class="inline">
-	<tr class="row0">
-		<td class="col0">Spot number</td><td class="col1">Sample</td>
-	</tr>
-	<tr class="row1">
-		<td class="col0">1.</td><td class="col1">GOPTS + nanobody + B52</td>
-	</tr>
-	<tr class="row2">
-		<td class="col0">2.</td><td class="col1">GOPTS + nanobody + B53</td>
-	</tr>
-	<tr class="row3">
-		<td class="col0">3.</td><td class="col1">GOPTS + nanobody + B54</td>
-	</tr>
-	<tr class="row4">
-		<td class="col0">4.</td><td class="col1">GOPTS + nanobody + B55</td>
-	</tr>
-	<tr class="row5">
-		<td class="col0">5.</td><td class="col1">GOPTS + nanobody + B56</td>
-	</tr>
-</table></div>
-<!-- EDIT3 TABLE [2479-2639] -->
-<p>
-<br/>
-</p>
-</div>
-<h4><a name="section07142016" id="section07142016">07/14/2016</a></h4>
-<div class="level4">
-<p>
-<strong>Sample preparation 1: GOPTS + anti-GFP nanobody + B52-B56</strong> <br/>
-</p>
-<ol>
-<li class="level1"><div class="li"> edges sealed with nail polish</div>
-</li>
-</ol>
-<p>
-<strong>Fluorescence microscopy 1 (Olympus 1&times;71): GOPTS + anti-GFP nanobody + B52-B56</strong>
-</p>
-<p>
-–&gt; pictures inconclusive:pictures inconclusive: wrong coverslips (second glass slide), no sufficient controls <br/>
-</p>
-<p>
-<strong>Sample preparation 2: GOPTS + anti-GFP nanobody + B52-56</strong><br/>
-</p>
-<ol>
-<li class="level1"><div class="li"> incubation with anti-GFP nanobody (5µl/spot, 330 µg/ml, 120 min) </div>
-</li>
-<li class="level1"><div class="li"> blocked with BSA(2%) (90 min)</div>
-</li>
-<li class="level1"><div class="li"> rinsed with PBS(1x)</div>
-</li>
-<li class="level1"><div class="li"> stored in PBS(1x) (for 9 days)</div>
-</li>
-</ol>
-<div class="table sectionedit4"><table class="inline">
-	<tr class="row0">
-		<td class="col0">Spot number</td><td class="col1">Sample</td>
-	</tr>
-	<tr class="row1">
-		<td class="col0">1.</td><td class="col1">GOPTS + nanobody + B52</td>
-	</tr>
-	<tr class="row2">
-		<td class="col0">2.</td><td class="col1">GOPTS + nanobody + B53</td>
-	</tr>
-	<tr class="row3">
-		<td class="col0">3.</td><td class="col1">GOPTS + nanobody + B54</td>
-	</tr>
-	<tr class="row4">
-		<td class="col0">4.</td><td class="col1">GOPTS + nanobody + B55</td>
-	</tr>
-	<tr class="row5">
-		<td class="col0">5.</td><td class="col1">GOPTS + nanobody + B56</td>
-	</tr>
-	<tr class="row6">
-		<td class="col0">6.</td><td class="col1">GOPTS + GFP (positive control)</td>
-	</tr>
-	<tr class="row7">
-		<td class="col0">7.</td><td class="col1">GOPTS + WT (negative control)</td>
-	</tr>
-</table></div>
-<!-- EDIT4 TABLE [3198-3429] -->
-<p>
-<br/>
-</p>
-</div>
-<h4><a name="section07152016" id="section07152016">07/15/2016</a></h4>
-<div class="level4">
-<p>
-<strong>Sample preparation</strong>
-</p>
-<ol>
-<li class="level1"><div class="li"> added nanobody to GOPTS plate (300 µg/ml, 5µl)</div>
-</li>
-<li class="level1"><div class="li"> tried to pipette rest nanobody down, but didn´t work</div>
-</li>
-<li class="level1"><div class="li"> blocked complete slide with BSA(2%)</div>
-</li>
-</ol>
-</div>
-<h4><a name="section0723016" id="section0723016">07/23/016</a></h4>
-<div class="level4">
-<p>
-<br/>
-<strong>Sample preparation 2: GOPTS + anti-GFP nanobody + B52-56</strong>
-</p>
-<ol>
-<li class="level1"><div class="li"> Washed slides with ddH2O</div>
-</li>
-<li class="level1"><div class="li"> Add spores (50µl/spot)(B52,B53,B54,B55,B65)</div>
-</li>
-</ol>
-<p>
-<br/>
-–&gt; FACS-analysis has shown that only B53 and B54 show fluorescence linked to GFP
-</p>
-<p>
-<strong>Coating of new slides with GOPTS (experiment 3)</strong>
-</p>
-<ol>
-<li class="level1"><div class="li"> according to protocol</div>
-</li>
-</ol>
-</div>
-<h4><a name="section07242016" id="section07242016">07/24/2016</a></h4>
-<div class="level4">
-<p>
-<strong>Preparation of samples 3: B53-B55</strong>
-</p>
-<ol>
-<li class="level1"><div class="li"> B53, B54, B55 (10µl/spot) on uncoated glass slide</div>
-</li>
-<li class="level1"><div class="li"> Mowiol DABCO (5µl/spot)</div>
-</li>
-<li class="level1"><div class="li"> covered with cover slip</div>
-</li>
-</ol>
-<div class="table sectionedit5"><table class="inline">
-	<tr class="row0">
-		<td class="col0">Spot number</td><td class="col1">Sample</td>
-	</tr>
-	<tr class="row1">
-		<td class="col0">1.</td><td class="col1">GOPTS + nanobody + B52</td>
-	</tr>
-	<tr class="row2">
-		<td class="col0">2.</td><td class="col1">GOPTS + nanobody + B53</td>
-	</tr>
-	<tr class="row3">
-		<td class="col0">3.</td><td class="col1">GOPTS + nanobody + B54</td>
-	</tr>
-</table></div>
-<!-- EDIT5 TABLE [4121-4225] -->
-<p>
-<br/>
-</p>
-<p>
-<strong>Fluorescence microscopy 2 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + B52-56</strong>
-</p>
-<ol>
-<li class="level1"><div class="li"> fluorescence signal of B53 and B54 </div>
-</li>
-</ol>
-<p>
-<strong>Fluorescence microscopy 3 (Olympus 1&times;71): B53-B55</strong>
-</p>
-<ol>
-<li class="level1"><div class="li"> fluorescence signal of positive controls (B53 and B54) but no signal of spores bound to GOPTS</div>
-</li>
-</ol>
-<p>
-B53 sample: no fluorescence observable (Air bubble)<br/>
-B54 sample: no fluorescence observable<br/>
-</p>
-<p>
-<strong>Sample preparation 4: GOPTS + anti-GFP nanobody + B53/B54</strong>
-</p>
-<ol>
-<li class="level1"><div class="li"> incubation with anti-GFP nanobody (5µl, 330µg/ml, 2d) </div>
-</li>
-</ol>
-</div>
-<h4><a name="section07262016" id="section07262016">07/26/2016</a></h4>
-<div class="level4">
-<p>
-<strong>Sample preparation 4: GOPTS + anti-GFP nanobody + B53/B54</strong>
-</p>
-<ol>
-<li class="level1"><div class="li"> Prepared new BSA solution with higher concentration (4% w/v)</div>
-</li>
-<li class="level1"><div class="li"> Blocked slides with BSA (4%) ,covered complete slide with BSA</div>
-</li>
-<li class="level1"><div class="li"> inkubated over night</div>
-</li>
-</ol>
-</div>
-<h4><a name="section07272016" id="section07272016">07/27/2016</a></h4>
-<div class="level4">
-<p>
-<strong>Sample preparation 4: GOPTS + anti-GFP nanobody + B53/B54</strong>
-</p>
-<ol>
-<li class="level1"><div class="li"> rinsed and washed with PBS(1x) (10 min, 10rpm)</div>
-</li>
-<li class="level1"><div class="li"> rinsed with water ddH2O</div>
-</li>
-<li class="level1"><div class="li"> dried with compressed air</div>
-</li>
-<li class="level1"><div class="li"> incubated with Mowiol DABCO (5µl/spot, overnight)</div>
-</li>
-</ol>
-<p>
-*Positive controls:
-</p>
-<ol>
-<li class="level1"><div class="li"> Slide with spores B53–&gt; fuorescence observable</div>
-</li>
-<li class="level1"><div class="li"> B54 –&gt;Fluorescence observable </div>
-</li>
-</ol>
-<div class="table sectionedit6"><table class="inline">
-	<tr class="row0">
-		<td class="col0">Spot number</td><td class="col1">Sample</td>
-	</tr>
-	<tr class="row1">
-		<td class="col0">1.</td><td class="col1">GOPTS + GFP (positive control)</td>
-	</tr>
-	<tr class="row2">
-		<td class="col0">2.</td><td class="col1">2	GOPTS + nanobody + GFP</td>
-	</tr>
-	<tr class="row3">
-		<td class="col0">3.</td><td class="col1">GOPTS + BSA + GFP (negative control)</td>
-	</tr>
-	<tr class="row4">
-		<td class="col0">4.</td><td class="col1">GOPTS + B53 (positive control)</td>
-	</tr>
-	<tr class="row5">
-		<td class="col0">5.</td><td class="col1">GOPTS + B54 (positive control)</td>
-	</tr>
-	<tr class="row6">
-		<td class="col0">6.</td><td class="col1">B53 (positive control)</td>
-	</tr>
-	<tr class="row7">
-		<td class="col0">7.</td><td class="col1">B54 (positive control)</td>
-	</tr>
-</table></div>
-<!-- EDIT6 TABLE [5340-5596] -->
-<p>
-<br/>
-</p>
-<p>
-<strong>Slides of experiment 3 </strong>
-</p>
-<ol>
-<li class="level1"><div class="li"> Washing with PBS</div>
-</li>
-<li class="level1"><div class="li"> rinsed with water ddH2O</div>
-</li>
-<li class="level1"><div class="li"> dried with compressed air</div>
-</li>
-</ol>
-<p>
-<strong>Positive controls exp. 3:</strong>
-</p>
-<ul>
-<li class="level1"><div class="li"> GFP direct on GOPTS</div>
-</li>
-<li class="level1"><div class="li"> GFP on nanobody</div>
-</li>
-<li class="level1"><div class="li"> GFP on BSA blocked surface</div>
-</li>
-<li class="level1"><div class="li"> Spores direct on GOPTS</div>
-</li>
-<li class="level1"><div class="li"> double concentrated spores direct on glass slide B53 and B54</div>
-</li>
-</ul>
-<p>
-Spores were added to glass surface and incubated until its dry
-</p>
-</div>
-<h4><a name="section07282016" id="section07282016">07/28/2016</a></h4>
-<div class="level4">
-<p>
-<strong>Fluorescence microscopy 4 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + B53/B54</strong>
-</p>
-<p>
-–&gt;spores appear to not have bound to nanobody
-</p>
-<ol>
-<li class="level1"><div class="li"> Washed slides of exp 3 with PBS</div>
-</li>
-<li class="level1"><div class="li"> added DABCO (5µl)</div>
-</li>
-</ol>
-</div>
-<h4><a name="section07292016" id="section07292016">07/29/2016</a></h4>
-<div class="level4">
-<p>
-<strong>prepared samples for fluorescence microscopy</strong>
-</p>
-</div>
-<h4><a name="section08082016" id="section08082016">08/08/2016</a></h4>
-<div class="level4">
-<p>
-<strong>Sample preparation 5:  GOPTS + anti-GFP nanobody + B53-56/GFP</strong>
-</p>
-<ol>
-<li class="level1"><div class="li"> incubation with anti-GFP nanobody (5µl, 330µg/ml, 60 min) </div>
-</li>
-<li class="level1"><div class="li"> rinsed and blocked with BSA(4%) (30 min)</div>
-</li>
-<li class="level1"><div class="li"> incubation with B53, B54, B55, B56, GFP (5µl/spot, 60 min)</div>
-</li>
-<li class="level1"><div class="li"> rinsed and washed with PBS(1x) (10 min, 10 rpm)</div>
-</li>
-<li class="level1"><div class="li"> rinsed with water ddH2O</div>
-</li>
-<li class="level1"><div class="li"> dried with compressed air</div>
-</li>
-<li class="level1"><div class="li"> incubated with Mowiol DABCO (5µl/spot, overnight)</div>
-</li>
-</ol>
-<div class="table sectionedit7"><table class="inline">
-	<tr class="row0">
-		<td class="col0">Spot number</td><td class="col1">Sample</td>
-	</tr>
-	<tr class="row1">
-		<td class="col0">1.</td><td class="col1">GOPTS + nanobody + B55 (negative control)</td>
-	</tr>
-	<tr class="row2">
-		<td class="col0">2.</td><td class="col1">GOPTS + B55 (negative control)</td>
-	</tr>
-	<tr class="row3">
-		<td class="col0">3.</td><td class="col1">GOPTS + nanobody + B53</td>
-	</tr>
-	<tr class="row4">
-		<td class="col0">4.</td><td class="col1">GOPTS + nanobody + GFP (positive control)</td>
-	</tr>
-	<tr class="row5">
-		<td class="col0">5.</td><td class="col1">GOPTS + BSA + GFP (negative control)</td>
-	</tr>
-	<tr class="row6">
-		<td class="col0">6.</td><td class="col1">B53 (positive control)</td>
-	</tr>
-	<tr class="row7">
-		<td class="col0">7.</td><td class="col1">B54 (positive control)</td>
-	</tr>
-	<tr class="row8">
-		<td class="col0">8.</td><td class="col1">GOPTS + GFP (positive control)</td>
-	</tr>
-	<tr class="row9">
-		<td class="col0">9.</td><td class="col1">GOPTS + B53 (positive control)</td>
-	</tr>
-	<tr class="row10">
-		<td class="col0">10.</td><td class="col1">GOPTS + nanobody(Alexa647) + B53</td>
-	</tr>
-	<tr class="row11">
-		<td class="col0">11.</td><td class="col1">GOPTS + nanobody(Alexa647) + B54</td>
-	</tr>
-	<tr class="row12">
-		<td class="col0">12.</td><td class="col1">GOPTS + nanobody(Alexa647) + WT(negative control)</td>
-	</tr>
-	<tr class="row13">
-		<td class="col0">13.</td><td class="col1">GOPTS + nanobody(Alexa647) + B56 (negative control)</td>
-	</tr>
-</table></div>
-<!-- EDIT7 TABLE [6696-7236] -->
-<p>
-<br/>
-</p>
-</div>
-<h4><a name="section08092016" id="section08092016">08/09/2016</a></h4>
-<div class="level4">
-<p>
-<strong>Test if spores bind Nanobody in solution</strong>
-</p>
-<ol>
-<li class="level1"><div class="li"> Wt, B53, B54, B56 centrifuged, resuspended in 50 µl nanobody-buffer, </div>
-</li>
-<li class="level1"><div class="li"> add 1µl labeled nanobody </div>
-</li>
-<li class="level1"><div class="li"> incubated 1h at 4°C</div>
-</li>
-<li class="level1"><div class="li"> washed 2 times 10 min with PBS</div>
-</li>
-</ol>
-<p>
-<strong>Problem:</strong> no introduction in the confocal microscopy mode
-</p>
-<ul>
-<li class="level1"><div class="li"> Flourescence microscopy </div>
-</li>
-</ul>
-<p>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:glass_b53_alexananobody_2016.08.09.jpg" class="media" title="labprotocol:glass_b53_alexananobody_2016.08.09.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:glass_b53_alexananobody_2016.08.09.jpg" class="mediacenter" alt="" width="200" /></a>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:gopts_nb_b55_wl.jpg" class="media" title="labprotocol:gopts_nb_b55_wl.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:gopts_nb_b55_wl.jpg" class="mediacenter" alt="" width="200" /></a>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:gopts_nb_b53_wl.jpg" class="media" title="labprotocol:gopts_nb_b53_wl.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:gopts_nb_b53_wl.jpg" class="mediacenter" alt="" width="200" /></a>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:gopts_gfp.jpg" class="media" title="labprotocol:gopts_gfp.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:gopts_gfp.jpg" class="mediacenter" alt="" width="200" /></a>
-</p>
-<p>
-<strong>Fluorescence microscopy 5 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + B53-56/GFP</strong>
-</p>
-<p>
-–&gt;only fluorescence detectable on slides 6 and 8
-</p>
-</div>
-<h4><a name="section08102016" id="section08102016">08/10/2016</a></h4>
-<div class="level4">
-<p>
-<strong>Sample preparation 6: anti-GFP nanobody + spores (WT, B53, B54, B56) (in solution)</strong>
-</p>
-<ol>
-<li class="level1"><div class="li"> incubation of anti-GFP nanobody (Alexa647-conjugated) (1µl, 330µg/ml, 60 min) + spores (WT, B53, B54, B56) (400µl, 60 min, 4°C) </div>
-</li>
-<li class="level1"><div class="li"> centrifuged (13400 rpm, 2 min)</div>
-</li>
-<li class="level1"><div class="li"> washed with PBS(1x) (1 ml)</div>
-</li>
-<li class="level1"><div class="li"> repeated steps 2 and 3</div>
-</li>
-<li class="level1"><div class="li"> centrifuged (13400 rpm, 2 min)</div>
-</li>
-<li class="level1"><div class="li"> resuspended in PBS(1x) (400µl)</div>
-</li>
-</ol>
-<div class="table sectionedit8"><table class="inline">
-	<tr class="row0">
-		<td class="col0">Spot number</td><td class="col1">Sample</td>
-	</tr>
-	<tr class="row1">
-		<td class="col0">1.</td><td class="col1">nanobody + WT (negative control)</td>
-	</tr>
-	<tr class="row2">
-		<td class="col0">2.</td><td class="col1">nanobody + B53</td>
-	</tr>
-	<tr class="row3">
-		<td class="col0">3.</td><td class="col1">nanobody + B53</td>
-	</tr>
-	<tr class="row4">
-		<td class="col0">4.</td><td class="col1">nanobody + B55 (negative control)</td>
-	</tr>
-	<tr class="row5">
-		<td class="col0">5.</td><td class="col1">nanobody + B56(negative control)</td>
-	</tr>
-	<tr class="row6">
-		<td class="col0">6.</td><td class="col1">nanobody + GFP (positive control)</td>
-	</tr>
-	<tr class="row7">
-		<td class="col0">7.</td><td class="col1">nanobody (positive control)</td>
-	</tr>
-	<tr class="row8">
-		<td class="col0">8.</td><td class="col1">GFP (positive control)</td>
-	</tr>
-</table></div>
-<!-- EDIT8 TABLE [8315-8590] -->
-<p>
-<br/>
-</p>
-<p>
-<strong>Fluorescence microscopy 6 (Nikon C2+ confocal microscope): anti-GFP nanobody + spores (WT, B53, B54, B56) in solution</strong>
-</p>
-<p>
-–&gt; no introduction to confocal mode: nanobody not observable
-</p>
-<p>
-<strong>Light microscopy</strong>
-Analyse the slides from 28.7.2016 under light mikroscope
-</p>
-</div>
-<h4><a name="section08142016" id="section08142016">08/14/2016</a></h4>
-<div class="level4">
-<p>
-<strong>Sample preparation 7: GOPTS + anti-GFP nanobody + B53/B54</strong>
-</p>
-<ol>
-<li class="level1"><div class="li"> incubated with nanobody (1µl, 330µg/ml, 60 min)</div>
-</li>
-<li class="level1"><div class="li"> rinsed and blocked with BSA(4%) (60 min)</div>
-</li>
-<li class="level1"><div class="li"> rinsed and washed twice with PBS(1x) (10 min, 10 rpm)</div>
-</li>
-<li class="level1"><div class="li"> rinsed with ddH2O </div>
-</li>
-<li class="level1"><div class="li"> dried with compressed air </div>
-</li>
-<li class="level1"><div class="li"> incubated with spores (10µl/spot, 120 min)</div>
-</li>
-<li class="level1"><div class="li"> rinsed and washed twice with PBS(1x) (10 min, 10rpm)</div>
-</li>
-<li class="level1"><div class="li"> rinsed with ddH20</div>
-</li>
-<li class="level1"><div class="li"> dried with compressed air </div>
-</li>
-<li class="level1"><div class="li"> incubated with Mowiol DABCO (5µl, overnight)</div>
-</li>
-</ol>
-<div class="table sectionedit9"><table class="inline">
-	<tr class="row0">
-		<td class="col0">Spot number</td><td class="col1">Sample</td>
-	</tr>
-	<tr class="row1">
-		<td class="col0">1.</td><td class="col1">nanobody + WT (negative control)</td>
-	</tr>
-	<tr class="row2">
-		<td class="col0">2.</td><td class="col1">nanobody + B53</td>
-	</tr>
-	<tr class="row3">
-		<td class="col0">3.</td><td class="col1">nanobody + B54</td>
-	</tr>
-	<tr class="row4">
-		<td class="col0">4.</td><td class="col1">nanobody + GFP (positive control)</td>
-	</tr>
-	<tr class="row5">
-		<td class="col0">5.</td><td class="col1">nanobody (positive control)</td>
-	</tr>
-	<tr class="row6">
-		<td class="col0">6.</td><td class="col1">GFP (positive control)</td>
-	</tr>
-</table></div>
-<!-- EDIT9 TABLE [9362-9560] -->
-<p>
-<br/>
-</p>
-<p>
-<strong>Washing test:</strong> We want to test if spores (B53) really bind to BSA surface like the previous test showed. We also want to test if the washing procedure has an impact on this result.
-</p>
-<ol>
-<li class="level1"><div class="li"> 5 slides coated with Gopts</div>
-</li>
-<li class="level1"><div class="li"> added 1 µl Nanobody (333µg/ml)</div>
-</li>
-<li class="level1"><div class="li"> incubated over night</div>
-</li>
-</ol>
-<p>
-<strong>Washing test A: GOPTS + anti-GFP nanobody + B53</strong><br/>
-</p>
-<ol>
-<li class="level1"><div class="li"> incubated with nanobody (1µl/mg, 300µg/ml, overnight, 4°C)</div>
-</li>
-</ol>
-</div>
-<h4><a name="section08152016" id="section08152016">08/15/2016</a></h4>
-<div class="level4">
-<p>
-<strong>Washing test A: GOPTS + anti-GFP nanobody + B53</strong><br/>
-</p>
-<p>
-<strong>Hier Bild von BSA + B53 mit GFP-signal</strong> <br/>
-<br/>
-</p>
-<ol>
-<li class="level1"><div class="li"> rinsed and  blocked with BSA(2%) (60 min, 10 rpm) </div>
-</li>
-<li class="level1"><div class="li"> rinsed with PBS(1x), washed PBS twice (10 min, 10 rpm)</div>
-</li>
-<li class="level1"><div class="li"> rinsed with ddH2O</div>
-</li>
-<li class="level1"><div class="li"> dried with compressed air </div>
-</li>
-<li class="level1"><div class="li"> incubated with B53 (10µl/spot, overnight)</div>
-</li>
-</ol>
-<p>
-<br/>
-</p>
-</div>
-<h4><a name="section08162016" id="section08162016">08/16/2016</a></h4>
-<div class="level4">
-<p>
-<strong>Washing test A: GOPTS + anti-GFP nanobody + B53</strong>
-</p>
-<p>
-<strong>conditions</strong> <br/>
-Coated plates were rinsed with PBS to remove the spore soluten. Slides were put in PBS in Petri dishes and put on the shaker.<br/>
-</p>
-<div class="table sectionedit10"><table class="inline">
-	<tr class="row0">
-		<td class="col0">Spot number</td><td class="col1">Sample</td><td class="col2">Condition</td>
-	</tr>
-	<tr class="row1">
-		<td class="col0">1.</td><td class="col1">GOPTS + nanobody + B53</td><td class="col2">washed 2x with PBS(1x) (10 min, 10 rpm)</td>
-	</tr>
-	<tr class="row2">
-		<td class="col0">2.</td><td class="col1">GOPTS + nanobody + B53</td><td class="col2">washed 3x with PBS(1x) (10 min, 10 rpm)</td>
-	</tr>
-	<tr class="row3">
-		<td class="col0">3.</td><td class="col1">GOPTS + nanobody + B53</td><td class="col2">washed 4x with PBS(1x) (10 min, 10 rpm)</td>
-	</tr>
-	<tr class="row4">
-		<td class="col0">4.</td><td class="col1">GOPTS + nanobody + B53</td><td class="col2">washed 1x with PBS(1x) + 0,05% Tween20 (10 min, 10 rpm)</td>
-	</tr>
-	<tr class="row5">
-		<td class="col0">5.</td><td class="col1">GOPTS + nanobody + B53</td><td class="col2">washed 1x with PBS(1x) + 0,1 % Tween20 (10 min, 10 rpm)</td>
-	</tr>
-</table></div>
-<!-- EDIT10 TABLE [10530-10932] -->
-<p>
-<br/>
-</p>
-</div>
-<h4><a name="section17082016" id="section17082016">17/08/2016</a></h4>
-<div class="level4">
-<p>
-<strong>Fluorescence microscopy 7 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + B53/B54</strong>
-</p>
-<p>
-–&gt; spores have not bound to nanobody<br/>
-</p>
-<p>
-<strong>Fluorescence microscopy A (Nikon C2+ confocal microscope): Washing test: GOPTS + anti-GFP nanobody + B53</strong>
-–&gt;spores have not bound to BSA, picture of BSA + B53 must have been a mistake<br/>
-</p>
-</div>
-<h4><a name="section08212016" id="section08212016">08/21/2016</a></h4>
-<div class="level4">
-<p>
-<strong>Sample preparation 8: GOPTS + anti-GFP nanobody + GFP</strong>
-</p>
-<p>
-–&gt;  10 slides activated with plasmagenerator coated with GOPTS.
-</p>
-<ol>
-<li class="level1"><div class="li"> incubated with nanobody (see table below, 60 min)</div>
-</li>
-<li class="level1"><div class="li"> rinsed and blocked with BSA(4%) (60 min)</div>
-</li>
-<li class="level1"><div class="li"> rinsed and washed with PBS(1x) (5 min, 10 rpm)</div>
-</li>
-<li class="level1"><div class="li"> rinsed with ddH2O </div>
-</li>
-<li class="level1"><div class="li"> dried with compressed air </div>
-</li>
-<li class="level1"><div class="li"> incubated with GFP (see table below, 1.35mg/ml, 20 min)</div>
-</li>
-<li class="level1"><div class="li"> rinsed and washed with PBS(1x) (10 min, 10rpm)</div>
-</li>
-<li class="level1"><div class="li"> rinsed with ddH20</div>
-</li>
-<li class="level1"><div class="li"> dried with compressed air </div>
-</li>
-<li class="level1"><div class="li"> incubated with Mowiol DABCO (5µl, overnight)<br/>
-</div>
-</li>
-</ol>
-<div class="table sectionedit11"><table class="inline">
-	<tr class="row0">
-		<td class="col0">Spot number</td><td class="col1">Sample</td>
-	</tr>
-	<tr class="row1">
-		<td class="col0">1.</td><td class="col1">GOPTS + GFP (1µl) (positive control)</td>
-	</tr>
-	<tr class="row2">
-		<td class="col0">2.</td><td class="col1">GOPTS + GFP (5 µl) (positive control)</td>
-	</tr>
-	<tr class="row3">
-		<td class="col0">3.</td><td class="col1">GOPTS + nanobody (1µl, 330µg/ml) + GFP</td>
-	</tr>
-	<tr class="row4">
-		<td class="col0">4.</td><td class="col1">GOPTS + nanobody (0.5µl, 10mg/ml) + GFP</td>
-	</tr>
-	<tr class="row5">
-		<td class="col0">5.</td><td class="col1">GOPTS + BSA + GFP (negative control)</td>
-	</tr>
-	<tr class="row6">
-		<td class="col0">6.</td><td class="col1">GOPTS + BSA + GFP (negative control)</td>
-	</tr>
-</table></div>
-<!-- EDIT11 TABLE [11856-12139] -->
-<p>
-<br/>
-</p>
-<p>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:freigem_1.5_2_10s_0_5ulnbunverduennt_.jpg" class="media" title="labprotocol:freigem_1.5_2_10s_0_5ulnbunverduennt_.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:freigem_1.5_2_10s_0_5ulnbunverduennt_.jpg" class="mediacenter" alt="" width="200" /></a><a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:freigem_2016_1.1_21.08.1ulgfpauf_gpots.jpg" class="media" title="labprotocol:freigem_2016_1.1_21.08.1ulgfpauf_gpots.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:freigem_2016_1.1_21.08.1ulgfpauf_gpots.jpg" class="mediacenter" alt="" width="200" /></a><a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:freigem_1.5_3_20s_0_5ulnbunverduennt_plusgfp.jpg" class="media" title="labprotocol:freigem_1.5_3_20s_0_5ulnbunverduennt_plusgfp.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:freigem_1.5_3_20s_0_5ulnbunverduennt_plusgfp.jpg" class="mediacenter" alt="" width="200" /></a>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:freigem_2016_1.2_10s_1ulnb1.30verdplus_gfp.jpg" class="media" title="labprotocol:freigem_2016_1.2_10s_1ulnb1.30verdplus_gfp.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:freigem_2016_1.2_10s_1ulnb1.30verdplus_gfp.jpg" class="mediacenter" alt="" width="200" /></a><a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:freigem_2016_1.4_10s5ulgfpaufgopts.jpg" class="media" title="labprotocol:freigem_2016_1.4_10s5ulgfpaufgopts.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:freigem_2016_1.4_10s5ulgfpaufgopts.jpg" class="mediacenter" alt="" width="200" /></a>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:freigem_2016_1.5_10s_0_5ulnbunverduennt.jpg" class="media" title="labprotocol:freigem_2016_1.5_10s_0_5ulnbunverduennt.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:freigem_2016_1.5_10s_0_5ulnbunverduennt.jpg" class="mediacenter" alt="" width="200" /></a>
-</p>
-<p>
-Picture after editing of shadow and contrast:
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:gfp2.png" class="media" title="labprotocol:gfp2.png"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:gfp2.png" class="mediacenter" alt="" width="200" /></a>
-Bedingungen: 5µl,GFP,20min. incubation, mit BSA blocking, 5min. waschen mit PBS,
-</p>
-<p>
-<strong>Blocking Test</strong>
-We want to test if the GOPTS is properly blocked by BSA and if milkprotein is also usefull for blocking
-</p>
-<p>
-Slide with :
-</p>
-<div class="table sectionedit12"><table class="inline">
-	<tr class="row0">
-		<td class="col0">GFP auf GOPTS</td><td class="col1" colspan="2">GFP good visible , border clearly visible</td>
-	</tr>
-	<tr class="row1">
-		<td class="col0">Milkprotein on GOPTS+GFP</td><td class="col1" colspan="2">No GFP observable</td>
-	</tr>
-	<tr class="row2">
-		<td class="col0">BSA on GOPTS + GFP</td><td class="col1" colspan="2">NO GFP observalbe</td>
-	</tr>
-</table></div>
-<!-- EDIT12 TABLE [12855-12999] -->
-<p>
-<strong>Contamination Test</strong>
-To avoid wrong interpretations of the fluorescence pictures we tested the chemikals and solutions we use, to find out if there are fluorescent contaminations.
-</p>
-<div class="table sectionedit13"><table class="inline">
-	<tr class="row0">
-		<td class="col0">solution</td><td class="col1">result (GFP LASER)</td>
-	</tr>
-	<tr class="row1">
-		<td class="col0">Mowiol/DABCO</td><td class="col1">a vew fluorecent crystals</td>
-	</tr>
-	<tr class="row2">
-		<td class="col0">PBS</td><td class="col1">a vew fluorecent crystals</td>
-	</tr>
-	<tr class="row3">
-		<td class="col0">BSA</td><td class="col1">no fluorescence</td>
-	</tr>
-</table></div>
-<!-- EDIT13 TABLE [13185-13309] -->
-<p>
-<br/>
-</p>
-</div>
-<h4><a name="section08222016" id="section08222016">08/22/2016</a></h4>
-<div class="level4">
-<p>
-<strong>Fluorescence microscopy 8 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + GFP</strong>
-<br/>
-</p>
-</div>
-<h4><a name="section08232016" id="section08232016">08/23/2016</a></h4>
-<div class="level4">
-<p>
-<strong>Sample preparation 9: GOPTS + GFP</strong>
-</p>
-<ol>
-<li class="level1"><div class="li"> incubated with GFP (1.35mg/ml, volume/incubation: see table below)</div>
-</li>
-<li class="level1"><div class="li"> rinsed and blocked with BSA(4%) (30 min)</div>
-</li>
-<li class="level1"><div class="li"> rinsed and washed with PBS(1x) (10 min, 10 rpm)</div>
-</li>
-<li class="level1"><div class="li"> rinsed with ddH2O </div>
-</li>
-<li class="level1"><div class="li"> dried with compressed air </div>
-</li>
-<li class="level1"><div class="li"> incubated with Mowiol DABCO (5µl, overnight)</div>
-</li>
-</ol>
-<div class="table sectionedit14"><table class="inline">
-	<tr class="row0">
-		<td class="col0">Spot number</td><td class="col1">Sample</td>
-	</tr>
-	<tr class="row1">
-		<td class="col0">1.</td><td class="col1">GOPTS + GFP (1 µl, 10 min) </td>
-	</tr>
-	<tr class="row2">
-		<td class="col0">2.</td><td class="col1">GOPTS + GFP (2 µl, 10 min) </td>
-	</tr>
-	<tr class="row3">
-		<td class="col0">3.</td><td class="col1">GOPTS + GFP (5 µl, 10 min)</td>
-	</tr>
-	<tr class="row4">
-		<td class="col0">4.</td><td class="col1">GOPTS + GFP (1 µl, 15 min)</td>
-	</tr>
-	<tr class="row5">
-		<td class="col0">5.</td><td class="col1">GOPTS + GFP (2 µl, 15 min)</td>
-	</tr>
-	<tr class="row6">
-		<td class="col0">6.</td><td class="col1">GOPTS + GFP (5 µl, 15 min)</td>
-	</tr>
-	<tr class="row7">
-		<td class="col0">7.</td><td class="col1">GOPTS + GFP (1 µl, 20 min)</td>
-	</tr>
-	<tr class="row8">
-		<td class="col0">8.</td><td class="col1">GOPTS + GFP (2 µl, 20 min)</td>
-	</tr>
-	<tr class="row9">
-		<td class="col0">9.</td><td class="col1">GOPTS + GFP (5 µl, 20 min)</td>
-	</tr>
-	<tr class="row10">
-		<td class="col0">10.</td><td class="col1">GOPTS (negative control)</td>
-	</tr>
-</table></div>
-<!-- EDIT14 TABLE [13756-14106] -->
-<p>
-<br/>
-<strong>Sample preparation 10: GOPTS + anti-GFP nanobody + - incubated with nanobody (1µl, 330µg/ml, 60 min)</strong>
-<br/>
-</p>
-<ol>
-<li class="level1"><div class="li"> rinsed and blocked with BSA(4%) (60 min)</div>
-</li>
-<li class="level1"><div class="li"> rinsed and washed with PBS(1x) (10 min, 10 rpm)</div>
-</li>
-<li class="level1"><div class="li"> rinsed with ddH2O </div>
-</li>
-<li class="level1"><div class="li"> dried with compressed air </div>
-</li>
-<li class="level1"><div class="li"> incubated with spores (10µl/spot, 30 min)</div>
-</li>
-<li class="level1"><div class="li"> rinsed and washed with PBS(1x) (10min, 10rpm)</div>
-</li>
-<li class="level1"><div class="li"> rinsed with ddH20</div>
-</li>
-<li class="level1"><div class="li"> dried with compressed air </div>
-</li>
-<li class="level1"><div class="li"> incubated with Mowiol DABCO (5µl, overnight)<br/>
-</div>
-</li>
-</ol>
-<p>
-<br/>
-</p>
-<div class="table sectionedit15"><table class="inline">
-	<tr class="row0">
-		<td class="col0">Spot number</td><td class="col1">Sample</td>
-	</tr>
-	<tr class="row1">
-		<td class="col0">1.</td><td class="col1">GOPTS + nanobody + B53</td>
-	</tr>
-	<tr class="row2">
-		<td class="col0">2.</td><td class="col1">GOPTS + nanobody + B53</td>
-	</tr>
-	<tr class="row3">
-		<td class="col0">3.</td><td class="col1">GOPTS + GFP(positive control)</td>
-	</tr>
-	<tr class="row4">
-		<td class="col0">4.</td><td class="col1">GOPTS + nanobody + GFP (positive control)</td>
-	</tr>
-	<tr class="row5">
-		<td class="col0">5.</td><td class="col1">GOPTS + BSA + B53 (negative control)</td>
-	</tr>
-</table></div>
-<!-- EDIT15 TABLE [14576-14776] -->
-<p>
-<br/>
-<strong>Washing test B: GOPTS + anti-GFP nanobody + B53</strong>
-</p>
-<ol>
-<li class="level1"><div class="li"> incubated with nanobody (1µl, 330µg/ml, 60 min)</div>
-</li>
-<li class="level1"><div class="li"> rinsed and blocked with BSA(4%) (60 min)</div>
-</li>
-<li class="level1"><div class="li"> rinsed and washed with PBS(1x) (10 min, 10 rpm)</div>
-</li>
-<li class="level1"><div class="li"> rinsed with ddH2O </div>
-</li>
-<li class="level1"><div class="li"> dried with compressed air </div>
-</li>
-<li class="level1"><div class="li"> incubated with spores (10µl/spot, 30 min)</div>
-</li>
-<li class="level1"><div class="li"> rinsed and washed with PBS(1x) (10 min, 10rpm)</div>
-</li>
-<li class="level1"><div class="li"> rinsed with ddH20</div>
-</li>
-<li class="level1"><div class="li"> dried with compressed air </div>
-</li>
-<li class="level1"><div class="li"> incubated with Mowiol DABCO (5µl, overnight)</div>
-</li>
-</ol>
-<p>
+<head>
-<br/>
+    <style>
+        .level3 {
+            font-size: 18px;
+        }
+        .pinkpurple {
+            background-color: #871150;
+            color: orange;
+        }
+        h3 {
+            font-size: 36px;
+            padding: 20px 0px;
+        }
+        strong {
+            color: grey;
+        }
+        .grey {
+            background-color: grey;
+            color: orange;
+        }
+        .image img {
+            margin: 20px;
+            width: 180px;
+        }
+        .image img:hover {
+            cursor: pointer;
+            -webkit-transform: scale(1.1);
+            /* Safari and Chrome */
+            -moz-transform: scale(1.1);
+            /* Firefox */
+            transition-duration: .2s;
+            transition-timing-function: linear;
+        }
+    </style>
+</head>
-</p>
+<body>
-<div class="table sectionedit16"><table class="inline">
+    <div class="color1">
-	<tr class="row0">
+        <div class="img_menu">
-		<td class="col0">Spot number</td><td class="col1">Sample</td><td class="col2">Condition</td>
+            <h5 style="text-align: center">Lab Journals </h5>
-	</tr>
+            <center>
-	<tr class="row1">
+                <div class="image">
-		<td class="col0">1.</td><td class="col1">GOPTS + nanobody + B53</td><td class="col2">rinsed with PBS(1x) </td>
+                    <a href="https://2016.igem.org/Team:Freiburg/NotebookCloning"> <img class="imga" src="https://static.igem.org/mediawiki/2016/1/14/T--Freiburg--CloningLab.png" alt="Smiley face"></a>
-	</tr>
+                    <a href="https://2016.igem.org/Team:Freiburg/NotebookSpores"> <img class="imgb" src="https://static.igem.org/mediawiki/2016/9/93/T--Freiburg--SporesLab.png" alt="Smiley face"></a>
-	<tr class="row2">
+                    <a href="https://2016.igem.org/Team:Freiburg/NotebookProteins"> <img class="imgc" src="https://static.igem.org/mediawiki/2016/2/23/T--Freiburg--ProteinsLab.png" alt="Smiley face"></a>
-		<td class="col0">2.</td><td class="col1">GOPTS + nanobody + B53</td><td class="col2">swivel with PBS(1x) (0.5 min)</td>
+                    <a href="https://2016.igem.org/Team:Freiburg/NotebookTargeting"><img class="imgd" src="https://static.igem.org/mediawiki/2016/f/f4/T--Freiburg--TargetingLab.png" alt="Smiley face"></a>
-	</tr>
+                    <a href="https://2016.igem.org/Team:Freiburg/NotebookDelivery"><img class="imge" src="https://static.igem.org/mediawiki/2016/4/4b/T--Freiburg--DeliveryLab.png" alt="Smiley face"></a>
-	<tr class="row3">
+                </div>
-		<td class="col0">3.</td><td class="col1">GOPTS + nanobody + B53</td><td class="col2">washed with PBS(1x) (1 min, 10 rpm)</td>
+            </center>
-	</tr>
+        </div>
-	<tr class="row4">
+    </div>
-		<td class="col0">4.</td><td class="col1">GOPTS + nanobody + B53</td><td class="col2">washed with PBS(1x) (2 min, 10 rpm)</td>
+    <div class="color8">
-	</tr>
+        <div class="para_center_20">
-	<tr class="row5">
+            <h5 class="sectionedit1">Group 4 - Adhesion</h5>
-		<td class="col0">5.</td><td class="col1">GOPTS + nanobody + B53</td><td class="col2">washed with PBS(1x) (3 min, 10 rpm)</td>
+            <div class="tags">
-	</tr>
+                The targeting is one of the main requirements the spores have to fulfill. An adhesion-assay was conducted to verify the binding of the spores to a desired target.
-	<tr class="row6">
+            </div>
-		<td class="col0">6.</td><td class="col1">GOPTS + nanobody + B53</td><td class="col2">washed with PBS(1x) (5 min, 10 rpm)</td>
+        </div>
-	</tr>
+        <!-- para_20-->
-	<tr class="row7">
+    </div>
-		<td class="col0">7.</td><td class="col1">GOPTS + BSA + B53(negative control)</td><td class="col2">washed with PBS(1x) (10 min, 10 rpm)</td>
+    <!-- color -->
-	</tr>
+    <div class="color2">
-	<tr class="row8">
+        <div class="para_20">
-		<td class="col0">8.</td><td class="col1">GOPTS + BSA + GFP(negative control)</td><td class="col2">washed with PBS(1x) (10 min, 10 rpm)</td>
+            <h3><a name="section07042016" id="section07042016">07/04/2016</a></h3>
-	</tr>
+            <div class="level4">
-	<tr class="row9">
+                <p> <strong>Bacillus subtilis strains used:</strong> </p>
-		<td class="col0">9.</td><td class="col1">GOPTS + GFP (positive control)</td><td class="col2">washed with PBS(1x) (10 min, 10 rpm)</td>
+                <div class="table sectionedit2">
-	</tr>
+                    <table class="inline">
-	<tr class="row10">
+                        <tr class="row0">
-		<td class="col0">10.</td><td class="col1">GOPTS + B53 (positive control)</td><td class="col2">washed with PBS(1x) (10 min, 10 rpm)</td>
+                            <td class="col0"><strong>Strain</strong></td>
-	</tr>
+                        </tr>
-</table></div>
+                        <tr class="row1">
-<!-- EDIT16 TABLE [15243-15939] -->
+                            <td class="col0">B52:B. Subtilis w168 amyE::PcgeA-CotZre-gfp-B0014(CM5)</td>
-<p>
+                        </tr>
-<br/>
+                        <tr class="row2">
+                            <td class="col0">B53:B. Subtilis w168 amyE::PcotYZ-CotZre-gfp-B0014(CM5)</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">B54:B. Subtilis w168 amyE::PcotYZ-CotZin-gfp-B0014(CM5)</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">B55:B. Subtilis w168 amyE::PcotV-CotZre-gfp-B0014(CM5)</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">B56:B. Subtilis w168 amyE::PCotv-CotZin-gfp-B0014(CM5)</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT2 TABLE [128-427] -->
+            </div>
+            <h3><a name="section07052016" id="section07052016">07/05/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Slide preparation: GOPTS</strong> </p>
+                <p> Surface activation of glass slides by plasma generator, coated with GOPTS 3-glycidyloxypropyl trimethoxysilane, washed with acetone and then stored for up to two weeeks. </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> labeled with a diamond pen</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> washed with EtOH then ddH2O</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried slides with compressed air</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> activation of surface with plasma generator</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with GOPTS (overnight)</div>
+                    </li>
+                </ol>
+            </div>
+            <h3><a name="section07102016" id="section07102016">07/10/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Buffer for anti-GFP nanobody (10 ml)</strong> </p>
+                <ul>
+                    <li class="level1">
+                        <div class="li"> 61mg Tris </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> 87mg NaCl </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> 1ml Glycerin </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> 2,5 ml Imidazole(1M)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> fill up with ddH2O to 10 ml</div>
+                    </li>
+                </ul>
+            </div>
+        </div>
+        <!-- para_20-->
+    </div>
+    <!-- color -->
+    <div class="color3">
+        <div class="para_20">
+            <h3><a name="section07122016" id="section07122016">07/12/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Sample preparation 1: GOPTS + anti-GFP nanobody + B52-B56</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubation with nanobody (0.5µl, 330µg/ml, overnight, 4°C) </div>
+                    </li>
+                </ol>
+            </div>
+            <h3><a name="section07132016" id="section07132016">07/13/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Sample preparation 1: GOPTS + anti-GFP nanobody + B52-B56</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> blocked with BSA(2%) (150µl/spot, 30 min) </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> washed with PBS(1x)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> washed with imidazole (20mM) in PBS(1x)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> washed with PBS(1x)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> washed with ddH2O</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with B52-B56 (100µl/spot, 120 min) </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with Mowiol DABCO(1,4-Diazabicyclo[2.2.2]octane) (5µl/spot, overnight), with second glass slide on top </div>
+                    </li>
+                </ol>
+                <div class="table sectionedit3">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1</td>
+                            <td class="col1">GOPTS + nanobody + B52</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">2</td>
+                            <td class="col1">GOPTS + nanobody + B53</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">3</td>
+                            <td class="col1">GOPTS + nanobody + B54</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">4</td>
+                            <td class="col1">GOPTS + nanobody + B55</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">5</td>
+                            <td class="col1">GOPTS + nanobody + B56</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT3 TABLE [1598-1758] -->
+            </div>
+            <h3><a name="section07142016" id="section07142016">07/14/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Sample preparation 1: GOPTS + anti-GFP nanobody + B52-B56</strong>
+                    <br/> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> edges sealed with nail polish</div>
+                    </li>
+                </ol>
+                <p> <strong>Fluorescence microscopy 1 (Olympus 1&times;71): GOPTS + anti-GFP nanobody + B52-B56</strong> </p>
+                <p> –&gt; pictures inconclusive: wrong coverslips (second glass slide), no sufficient controls
+                    <br/> </p>
+                <p> <strong>Sample preparation 2: GOPTS + anti-GFP nanobody + B52-56</strong>
+                    <br/> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubation with anti-GFP nanobody (5µl/spot, 330 µg/ml, 120 min) </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> blocked with BSA(2%) (90 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with PBS(1x)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> stored in PBS(1x) (for 9 days)</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit4">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1</td>
+                            <td class="col1">GOPTS + nanobody + B52</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">2</td>
+                            <td class="col1">GOPTS + nanobody + B53</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">3</td>
+                            <td class="col1">GOPTS + nanobody + B54</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">4</td>
+                            <td class="col1">GOPTS + nanobody + B55</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">5</td>
+                            <td class="col1">GOPTS + nanobody + B56</td>
+                        </tr>
+                        <tr class="row6">
+                            <td class="col0">6</td>
+                            <td class="col1">GOPTS + GFP (positive control)</td>
+                        </tr>
+                        <tr class="row7">
+                            <td class="col0">7</td>
+                            <td class="col1">GOPTS + WT (negative control)</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT4 TABLE [2287-2516] -->
+            </div>
+            <h3><a name="section07152016" id="section07152016">07/15/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Sample preparation</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> added nanobody to GOPTS plate (300 µg/ml, 5µl)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> tried to pipette rest nanobody down, but didn´t work</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> blocked complete slide with BSA(2%)</div>
+                    </li>
+                </ol>
+            </div>
+        </div>
+        <!-- para_20-->
+    </div>
+    <!-- color -->
+    <div class="color4">
+        <div class="para_20">
+            <h3><a name="section0723016" id="section0723016">07/23/016</a></h3>
+            <div class="level4">
+                <p> <strong>Sample preparation 2: GOPTS + anti-GFP nanobody + B52-56</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> washed slides with ddH2O</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with B52, B53, B54, B55, B56 (50µl/spot, overnight)</div>
+                    </li>
+                </ol>
+                <p> –&gt; <strong><em class="u">FACS-analysis has shown that only B53 and B54 show fluorescence linked to GFP</em></strong> </p>
+            </div>
+            <h3><a name="section07242016" id="section07242016">07/24/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Preparation of samples 3: B53-B55</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> B53, B54, B55 (10µl/spot) on uncoated glass slide</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> Mowiol DABCO (5µl/spot)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> covered with cover slip</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit5">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1</td>
+                            <td class="col1">B52</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">2</td>
+                            <td class="col1">B53</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">3</td>
+                            <td class="col1">B54</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT5 TABLE [3152-3201] -->
+                <p> <strong>Fluorescence microscopy 2 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + B52-56</strong> </p>
+                <p> –&gt; fluorescence signal of B53 and B54 </p>
+                <p> <strong>Fluorescence microscopy 3 (Olympus 1&times;71): B53-B55</strong> </p>
+                <p> –&gt; fluorescence signal of positive controls (B53 and B54) but no signal of spores bound to GOPTS </p>
+                <p> <strong>Sample preparation 4: GOPTS + anti-GFP nanobody + B53/B54</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubation with anti-GFP nanobody (5µl, 330µg/ml, 2d) </div>
+                    </li>
+                </ol>
+            </div>
+            <h3><a name="section07262016" id="section07262016">07/26/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Sample preparation 4: GOPTS + anti-GFP nanobody + B53/B54</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> blocked with BSA(4%) (overnight)</div>
+                    </li>
+                </ol>
+            </div>
+        </div>
+        <!-- para_20-->
+    </div>
+    <!-- color -->
+    <div class="color5">
+        <div class="para_20">
+            <h3><a name="section07272016" id="section07272016">07/27/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Sample preparation 4: GOPTS + anti-GFP nanobody + B53/B54</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (10 min, 10rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with water ddH2O</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with Mowiol DABCO (5µl/spot, overnight)</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit6">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1</td>
+                            <td class="col1">GOPTS + GFP (positive control)</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">2</td>
+                            <td class="col1">GOPTS + nanobody + GFP</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">3</td>
+                            <td class="col1">GOPTS + BSA + GFP (negative control)</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">4</td>
+                            <td class="col1">GOPTS + B53 (positive control)</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">5</td>
+                            <td class="col1">GOPTS + B54 (positive control)</td>
+                        </tr>
+                        <tr class="row6">
+                            <td class="col0">6</td>
+                            <td class="col1">B53 (positive control)</td>
+                        </tr>
+                        <tr class="row7">
+                            <td class="col0">7</td>
+                            <td class="col1">B54 (positive control)</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT6 TABLE [3988-4240] -->
+            </div>
+            <h3><a name="section07282016" id="section07282016">07/28/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Fluorescence microscopy 4 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + B53/B54</strong> </p>
+                <p> –&gt; spores appear to not have been bound to nanobody </p>
+            </div>
+            <h3><a name="section08082016" id="section08082016">08/08/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Sample preparation 5:  GOPTS + anti-GFP nanobody + B53-56/GFP</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubation with anti-GFP nanobody (5µl, 330µg/ml, 60 min) </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and blocked with BSA(4%) (30 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with B53, B54, B55, B56, GFP (5µl/spot, 60 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (10 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with water ddH2O</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with Mowiol DABCO (5µl/spot, overnight)</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit7">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1</td>
+                            <td class="col1">GOPTS + nanobody + B55 (negative control)</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">2</td>
+                            <td class="col1">GOPTS + B55 (negative control)</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">3</td>
+                            <td class="col1">GOPTS + nanobody + B53</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">4</td>
+                            <td class="col1">GOPTS + nanobody + GFP (positive control)</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">5</td>
+                            <td class="col1">GOPTS + BSA + GFP (negative control)</td>
+                        </tr>
+                        <tr class="row6">
+                            <td class="col0">6</td>
+                            <td class="col1">B53 (positive control)</td>
+                        </tr>
+                        <tr class="row7">
+                            <td class="col0">7</td>
+                            <td class="col1">B54 (positive control)</td>
+                        </tr>
+                        <tr class="row8">
+                            <td class="col0">8</td>
+                            <td class="col1">GOPTS + GFP (positive control)</td>
+                        </tr>
+                        <tr class="row9">
+                            <td class="col0">9</td>
+                            <td class="col1">GOPTS + B53 (positive control)</td>
+                        </tr>
+                        <tr class="row10">
+                            <td class="col0">10</td>
+                            <td class="col1">GOPTS + nanobody(Alexa647) + B53</td>
+                        </tr>
+                        <tr class="row11">
+                            <td class="col0">11</td>
+                            <td class="col1">GOPTS + nanobody(Alexa647) + B54</td>
+                        </tr>
+                        <tr class="row12">
+                            <td class="col0">12</td>
+                            <td class="col1">GOPTS + nanobody(Alexa647) + WT(negative control)</td>
+                        </tr>
+                        <tr class="row13">
+                            <td class="col0">13</td>
+                            <td class="col1">GOPTS + nanobody(Alexa647) + B56 (negative control)</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT7 TABLE [4840-5372] -->
+            </div>
+        </div>
+        <!-- para_20-->
+    </div>
+    <!-- color -->
+    <div class="color6">
+        <div class="para_20">
+            <h3><a name="section08092016" id="section08092016">08/09/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Fluorescence microscopy 5 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + B53-56/GFP</strong> </p>
+                <p> –&gt; spores have not been bound </p>
+            </div>
+            <h3><a name="section08102016" id="section08102016">08/10/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Sample preparation 6: anti-GFP nanobody + spores (WT, B53, B54, B56) (in solution)</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubation of anti-GFP nanobody (Alexa647-conjugated) (1µl, 330µg/ml, 60 min) + spores (WT, B53, B54, B56) (400µl, 60 min, 4°C) </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> centrifuged (13400 rpm, 2 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> washed with PBS(1x) (1 ml)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> repeated steps 2 and 3</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> centrifuged (13400 rpm, 2 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> resuspended in PBS(1x) (400µl)</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit8">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1</td>
+                            <td class="col1">nanobody + WT (negative control)</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">2</td>
+                            <td class="col1">nanobody + B53</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">3</td>
+                            <td class="col1">nanobody + B53</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">4</td>
+                            <td class="col1">nanobody + B55 (negative control)</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">5</td>
+                            <td class="col1">nanobody + B56(negative control)</td>
+                        </tr>
+                        <tr class="row6">
+                            <td class="col0">6</td>
+                            <td class="col1">nanobody + GFP (positive control)</td>
+                        </tr>
+                        <tr class="row7">
+                            <td class="col0">7</td>
+                            <td class="col1">nanobody (positive control)</td>
+                        </tr>
+                        <tr class="row8">
+                            <td class="col0">8</td>
+                            <td class="col1">GFP (positive control)</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT8 TABLE [5936-6208] -->
+                <p> <strong>Fluorescence microscopy 6 (Nikon C2+ confocal microscope): anti-GFP nanobody + spores (WT, B53, B54, B56) in solution</strong> </p>
+                <p> –&gt; no introduction to confocal mode: nanobody not observable </p>
+            </div>
+            <h3><a name="section08142016" id="section08142016">08/14/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Sample preparation 7: GOPTS + anti-GFP nanobody + B53/B54</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubated with nanobody (1µl, 330µg/ml, 60 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and blocked with BSA(4%) (60 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed twice with PBS(1x) (10 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH2O </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubated with spores (10µl/spot, 120 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed twice with PBS(1x) (10 min, 10rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH20</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with Mowiol DABCO (5µl, overnight)</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit9">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1</td>
+                            <td class="col1">nanobody + WT (negative control)</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">2</td>
+                            <td class="col1">nanobody + B53</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">3</td>
+                            <td class="col1">nanobody + B54</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">4</td>
+                            <td class="col1">nanobody + GFP (positive control)</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">5</td>
+                            <td class="col1">nanobody (positive control)</td>
+                        </tr>
+                        <tr class="row6">
+                            <td class="col0">6</td>
+                            <td class="col1">GFP (positive control)</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT9 TABLE [6899-7096] -->
+                <p> <strong> Washing test A: GOPTS + anti-GFP nanobody + B53</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubation with nanobody (1µl/mg, 300µg/ml, overnight, 4°C)</div>
+                    </li>
+                </ol>
+            </div>
+        </div>
+        <!-- para_20-->
+    </div>
+    <!-- color -->
+    <div class="color7">
+        <div class="para_20">
+            <h3><a name="section08152016" id="section08152016">08/15/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Washing test A: GOPTS + anti-GFP nanobody + B53</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> rinsed and blocked with BSA(2%) (60 min, 10 rpm) </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with PBS(1x), washed PBS twice (10 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH2O</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with B53 (10µl/spot, overnight)</div>
+                    </li>
+                </ol>
+            </div>
+            <h3><a name="section08162016" id="section08162016">08/16/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Washing test A: GOPTS + anti-GFP nanobody + B53</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed (see table below)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddh2O</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubated with Mowiol DABCO (5µl, overnight)</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit10">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                            <td class="col2"><strong>Condition</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1</td>
+                            <td class="col1">GOPTS + nanobody + B53</td>
+                            <td class="col2">washed 2x with PBS(1x) (10 min, 10 rpm)</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">2</td>
+                            <td class="col1">GOPTS + nanobody + B53</td>
+                            <td class="col2">washed 3x with PBS(1x) (10 min, 10 rpm)</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">3</td>
+                            <td class="col1">GOPTS + nanobody + B53</td>
+                            <td class="col2">washed 4x with PBS(1x) (10 min, 10 rpm)</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">4</td>
+                            <td class="col1">GOPTS + nanobody + B53</td>
+                            <td class="col2">washed 1x with PBS(1x) + 0,05% Tween20 (10 min, 10 rpm)</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">5</td>
+                            <td class="col1">GOPTS + nanobody + B53</td>
+                            <td class="col2">washed 1x with PBS(1x) + 0,1 % Tween20 (10 min, 10 rpm)</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT10 TABLE [7723-8129] -->
+            </div>
+            <h3><a name="section08172016" id="section08172016">08/17/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Fluorescence microscopy 7 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + B53/B54</strong> </p>
+                <p> –&gt; spores have not bound to nanobody </p>
+                <p> <strong>Fluorescence microscopy A (Nikon C2+ confocal microscope): Washing test: GOPTS + anti-GFP nanobody + B53</strong> </p>
+                <p> –&gt; spores have not bound to BSA </p>
+            </div>
+        </div>
+        <!-- para_20-->
+    </div>
+    <!-- color -->
+    <div class="color8">
+        <div class="para_20">
+            <h3><a name="section08212016" id="section08212016">08/21/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Sample preparation 8: GOPTS + anti-GFP nanobody + GFP</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubation with nanobody (see table below, 60 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and blocked with BSA(4%) (60 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (5 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH2O </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubated with GFP (see table below, 1.35mg/ml, 20 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (10 min, 10rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH20</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with Mowiol DABCO (5µl, overnight)
+                            <br/> </div>
+                    </li>
+                </ol>
+                <div class="table sectionedit11">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1</td>
+                            <td class="col1">GOPTS + GFP (1µl) (positive control)</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">2</td>
+                            <td class="col1">GOPTS + GFP (5 µl) (positive control)</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">3</td>
+                            <td class="col1">GOPTS + nanobody (1µl, 330µg/ml) + GFP</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">4</td>
+                            <td class="col1">GOPTS + nanobody (0.5µl, 10mg/ml) + GFP</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">5, 6</td>
+                            <td class="col1">GOPTS + BSA + GFP (negative control)</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT11 TABLE [8932-9176] -->
+            </div>
+            <h3><a name="section08222016" id="section08222016">08/22/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Fluorescence microscopy 8 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + GFP</strong> </p>
+                <p> –&gt; GFP not bound, GOPTS plates too old </p>
+            </div>
+            <h3><a name="section08232016" id="section08232016">08/23/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Sample preparation 9: GOPTS + GFP</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubation with GFP (1.35mg/ml, volume/incubation: see table below)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and blocked with BSA(4%) (30 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (10 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH2O </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with Mowiol DABCO (5µl, overnight)</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit12">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1</td>
+                            <td class="col1">GOPTS + GFP (1 µl, 10 min)</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">2</td>
+                            <td class="col1">GOPTS + GFP (2 µl, 10 min)</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">3</td>
+                            <td class="col1">GOPTS + GFP (5 µl, 10 min)</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">4</td>
+                            <td class="col1">GOPTS + GFP (1 µl, 15 min)</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">5</td>
+                            <td class="col1">GOPTS + GFP (2 µl, 15 min)</td>
+                        </tr>
+                        <tr class="row6">
+                            <td class="col0">6</td>
+                            <td class="col1">GOPTS + GFP (5 µl, 15 min)</td>
+                        </tr>
+                        <tr class="row7">
+                            <td class="col0">7</td>
+                            <td class="col1">GOPTS + GFP (1 µl, 20 min)</td>
+                        </tr>
+                        <tr class="row8">
+                            <td class="col0">8</td>
+                            <td class="col1">GOPTS + GFP (2 µl, 20 min)</td>
+                        </tr>
+                        <tr class="row9">
+                            <td class="col0">9</td>
+                            <td class="col1">GOPTS + GFP (5 µl, 20 min)</td>
+                        </tr>
+                        <tr class="row10">
+                            <td class="col0">10</td>
+                            <td class="col1">GOPTS (negative control)</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT12 TABLE [9664-10007] -->
+                <p> <strong>Sample preparation 10: GOPTS + anti-GFP nanobody + B53</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubation with nanobody (1µl, 330µg/ml, 60 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and blocked with BSA(4%) (60 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (10 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH2O </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubated with spores (10µl/spot, 30 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (10min, 10rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH20</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with Mowiol DABCO (5µl, overnight)</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit13">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1</td>
+                            <td class="col1">GOPTS + nanobody + B53</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">2</td>
+                            <td class="col1">GOPTS + nanobody + B53</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">3</td>
+                            <td class="col1">GOPTS + GFP(positive control)</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">4</td>
+                            <td class="col1">GOPTS + nanobody + GFP (positive control)</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">5</td>
+                            <td class="col1">GOPTS + BSA + B53 (negative control)</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT13 TABLE [10477-10677] -->
+                <p> <strong>Washing test B: GOPTS + anti-GFP nanobody + B53</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubation with nanobody (1µl, 330µg/ml, 60 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and blocked with BSA(4%) (60 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (10 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH2O </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubated with spores (10µl/spot, 30 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (10 min, 10rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH20</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with Mowiol DABCO (5µl, overnight)</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit14">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                            <td class="col2"><strong>Condition</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1</td>
+                            <td class="col1">GOPTS + nanobody + B53</td>
+                            <td class="col2">rinsed with PBS(1x) </td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">2</td>
+                            <td class="col1">GOPTS + nanobody + B53</td>
+                            <td class="col2">swivel with PBS(1x) (0.5 min)</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">3</td>
+                            <td class="col1">GOPTS + nanobody + B53</td>
+                            <td class="col2">washed with PBS(1x) (1 min, 10 rpm)</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">4</td>
+                            <td class="col1">GOPTS + nanobody + B53</td>
+                            <td class="col2">washed with PBS(1x) (2 min, 10 rpm)</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">5</td>
+                            <td class="col1">GOPTS + nanobody + B53</td>
+                            <td class="col2">washed with PBS(1x) (3 min, 10 rpm)</td>
+                        </tr>
+                        <tr class="row6">
+                            <td class="col0">6</td>
+                            <td class="col1">GOPTS + nanobody + B53</td>
+                            <td class="col2">washed with PBS(1x) (5 min, 10 rpm)</td>
+                        </tr>
+                        <tr class="row7">
+                            <td class="col0">7</td>
+                            <td class="col1">GOPTS + BSA + B53(negative control)</td>
+                            <td class="col2">washed with PBS(1x) (10 min, 10 rpm)</td>
+                        </tr>
+                        <tr class="row8">
+                            <td class="col0">8</td>
+                            <td class="col1">GOPTS + BSA + GFP(negative control)</td>
+                            <td class="col2">washed with PBS(1x) (10 min, 10 rpm)</td>
+                        </tr>
+                        <tr class="row9">
+                            <td class="col0">9</td>
+                            <td class="col1">GOPTS + GFP (positive control)</td>
+                            <td class="col2">washed with PBS(1x) (10 min, 10 rpm)</td>
+                        </tr>
+                        <tr class="row10">
+                            <td class="col0">10</td>
+                            <td class="col1">GOPTS + B53 (positive control)</td>
+                            <td class="col2">washed with PBS(1x) (10 min, 10 rpm)</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT14 TABLE [11141-11836] -->
+            </div>
+        </div>
+        <!-- para_20-->
+    </div>
+    <!-- color -->
+    <div class="color1">
+        <div class="para_20">
+            <h3><a name="section08242016" id="section08242016">08/24/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Fluorescence microscopy 9 (Nikon C2+ confocal microscope): GOPTS + GFP</strong> </p>
+                <p> –&gt; strong fluorescence signal at some of the edges: incubation too long/volume too little, GFP has dried and could not have been washed away properly </p>
+                <p>
+                  <center><img class="something" src="https://static.igem.org/mediawiki/2016/5/5c/T--Freiburg--targetingjournal1.png"> </center>
+                  <center><img class="something" src="https://static.igem.org/mediawiki/2016/8/87/T--Freiburg--targetingjournal2.png"> </center>
+                  <center><img class="something" src="https://static.igem.org/mediawiki/2016/2/25/T--Freiburg--targetingjournal3.png"> </center>
+                  <center><img class="something" src="https://static.igem.org/mediawiki/2016/6/66/T--Freiburg--targetingjournal4.png"> </center>
+                  <center><img class="something" src="https://static.igem.org/mediawiki/2016/4/46/T--Freiburg--targetingjournal5.png"> </center>
+                  <center><img class="something" src="https://static.igem.org/mediawiki/2016/0/00/T--Freiburg--targetingjournal6.png"> </center>
+                  <center><img class="something" src="https://static.igem.org/mediawiki/2016/a/af/T--Freiburg--targetingjournal7.png"> </center>
-</p>
+                </p>
+                <ul>
+                    <li class="level1">
+                        <div class="li"> 1) GOPTS + GFP (1 µl, 10 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> 2) GOPTS + GFP (2 µl, 10 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> 3) GOPTS + GFP (5 µl, 10 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> 4) GOPTS + GFP (1 µl, 15 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> 5) GOPTS + GFP (2 µl, 15 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> 6) GOPTS + GFP (5 µl, 15 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> 7) GOPTS + GFP (1 µl, 20 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> 8) GOPTS + GFP (2 µl, 20 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> 9) GOPTS + GFP (5 µl, 20 min)</div>
+                    </li>
+                </ul>
+                <p> <strong>Fluorescence microscopy 10 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + B53</strong> </p>
+                <p> –&gt; no spores appear to have bound to nanobody </p>
+                <p> <strong>Fluorescence microscopy B (Nikon C2+ confocal microscope): Washing test: GOPTS + anti-GFP nanobody + B53</strong> </p>
+                <p> –&gt; no spores after washing, no binding between nanobody and B53? </p>
+            </div>
+            <h3><a name="section08252016" id="section08252016">08/25/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Blocking test C: GOPTS + BSA/milk powder + GFP</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> blocking with BSA(4%)/milk powder(5%) (120 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (10 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with GFP (1µl/spot, 1.35mg/ml, 20 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (10 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH20</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with Mowiol DABCO (5µl, overnight)</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit15">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1</td>
+                            <td class="col1">GOPTS + GFP (positive control)</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">2</td>
+                            <td class="col1">GOPTS + milk powder(5%) + GFP</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">3</td>
+                            <td class="col1">GOPTS + BSA(4%) + GFP</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT15 TABLE [13676-13797] -->
+                <p> <strong>Contamination Test D: BSA, PBS Mowiol DABCO</strong> </p>
+                <p> –&gt; no contamination/pollution detected </p>
+                <p> <strong>Sample preparation 11: GOPTS + mCherry</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubated with mCherry (1µl, 2mg/ml, incubation: see table below)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and blocked with BSA(4%) (30 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (10 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH2O </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubated with Mowiol DABCO (5µl, overnight)</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit16">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1</td>
+                            <td class="col1">GOPTS + mCherry (5 min) </td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">2</td>
+                            <td class="col1">GOPTS + mCherry (10 min)</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">3</td>
+                            <td class="col1">GOPTS + mCherry (15 min)</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">4</td>
+                            <td class="col1">GOPTS (negative control)</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">5</td>
+                            <td class="col1">mCherry + Mowiol DABCO (positive control)</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT16 TABLE [14206-14394] -->
+                <p> <strong>Sample preparation 12: GOPTS + anti-GFP nanobody + GFP</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubated with nanobody (5µl, 330µg/ml, 120 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and blocked with milk powder(5%) (60 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (10 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH2O </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubated with GFP (2µl, 1.35mg/ml, 20 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (10 min, 10rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH20</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubated with Mowiol DABCO (5µl, overnight)</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit17">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1</td>
+                            <td class="col1">GOPTS + nanobody + GFP</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">2</td>
+                            <td class="col1">GOPTS (negative control)</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">3</td>
+                            <td class="col1">GOPTS + GFP (positive control)</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">4</td>
+                            <td class="col1">GOPTS + nanobody (negative control)</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">5</td>
+                            <td class="col1">GOPTS + BSA + GFP (negative control)</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT17 TABLE [14873-15071] -->
+            </div>
+        </div>
+        <!-- para_20-->
+    </div>
+    <!-- color -->
+    <div class="color2">
+        <div class="para_20">
+            <h3><a name="section08272016" id="section08272016">08/27/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Fluorescence microscopy C (Nikon C2+ confocal microscope): Blocking test: GOPTS + BSA/milkpowder + GFP</strong> </p>
+                <p> –&gt; No GFP signal after blocking with BSA and milk powder </p>
+                <p> <strong>Fluorescence microscopy 11 (Nikon C2+ confocal microscope): GOPTS + mCherry</strong> </p>
+                <p> –&gt; no mCherry-signal, too weak (quantum yield of mCherry about 1/3 of that of GFP) </p>
+                <p> <strong>Fluorescence microscopy 12 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + GFP</strong> </p>
+                <p> –&gt; GFP has not bound to nanobody </p>
+            </div>
+            <h3><a name="section08302016" id="section08302016">08/30/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Sample preparation 13: GOPTS + nanobody + GFP</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubated with nanobody (5µl, 330µg/ml, 60 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and blocked with milk powder(5%) (30 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (10 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH2O </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubated with GFP (2µl, 1.35mg/ml, 20 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (10 min, 10rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH20</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubated with Mowiol DABCO (5µl, overnight)</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit18">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1,2</td>
+                            <td class="col1">GOPTS + nanobody(Alexa647) + GFP</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">3,4</td>
+                            <td class="col1">GOPTS + nanobody + GFP</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">5</td>
+                            <td class="col1">GOPTS + GFP (positive control)</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">6</td>
+                            <td class="col1">GOPTS + nanobody (negative control)</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">7</td>
+                            <td class="col1">GOPTS + BSA (negative control)</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT18 TABLE [16042-16246] -->
+            </div>
+            <h3><a name="section09012016" id="section09012016">09/01/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Fluorescence microscopy 13 (Nikon C2+ confocal microscope): GOPTS + nanobody + GFP</strong> </p>
+                <p> –&gt; no GFP signal detectable </p>
+            </div>
+        </div>
+        <!-- para_20-->
+    </div>
+    <!-- color -->
+    <div class="color3">
+        <div class="para_20">
+            <h3><a name="section09062016" id="section09062016">09/06/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Sample preparation 14: GOPTS + GFP</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubated with GFP (10µl, 1.35mg/ml, 30 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and blocked with milk powder(5%) (30 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (10 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH2O </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubated with Mowiol DABCO (5µl, overnight)</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit19">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1,2</td>
+                            <td class="col1">GOPTS + mCherry</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">3</td>
+                            <td class="col1">GOPTS + BSA + mCherry (negative control)</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">4</td>
+                            <td class="col1">GOPTS + BSA (negative control)</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">5</td>
+                            <td class="col1">mCherry (positive control)</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT19 TABLE [16703-16861] -->
+                <p> <em class="u">Mowiol was produced due to this protocol:</em> <a href="https://static.igem.org/mediawiki/2016/5/56/T--Freiburg--LabJournal98.pdf" class="urlextern" target="_Blank" title="https://www.carlroth.com/downloads/ba/en/0/BA_0713_EN.pdf" rel="nofollow">https://www.carlroth.com/downloads/ba/en/0/BA_0713_EN.pdf</a>
+                    <br/> </p>
+            </div>
+            <h3><a name="section09072016" id="section09072016">09/07/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Fluorescence microscopy 14 (Nikon C2+ confocal microscope): GOPTS + GFP</strong> </p>
+                <p> –&gt; spots with evenly distributed GFP </p>
+                <p>
+                    <center><img class="something" src="https://static.igem.org/mediawiki/2016/c/cc/T--Freiburg--targetingjournal8.png"> </center>
+                  <center><img class="something" src="https://static.igem.org/mediawiki/2016/a/a1/T--Freiburg--targetingjournal9.png"> </center>
+                    <br/> </p>
+                <ul>
+                    <li class="level1">
+                        <div class="li"> 1) GOPTS + GFP</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> 2) GOPTS + nanobody + GFP</div>
+                    </li>
+                </ul>
+                <p>
+                    <br/> </p>
+            </div>
+            <h3><a name="section09142016" id="section09142016">09/14/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Sample preparation 16: GOPTS + B53</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubated with B53 (5µl, 60 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and blocked with milk powder(4%) (30 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (10 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH2O </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubated with Mowiol DABCO (5µl, overnight)</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit20">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1,2,3</td>
+                            <td class="col1">GOPTS + B53</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">4</td>
+                            <td class="col1">GOPTS + WT (negative control)</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">5</td>
+                            <td class="col1">WT (negative control)</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">6</td>
+                            <td class="col1">B53 (positive control)</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">7</td>
+                            <td class="col1">GOPTS + GFP</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT20 TABLE [17589-17737] -->
+            </div>
+        </div>
+        <!-- para_20-->
+    </div>
+    <!-- color -->
+    <div class="color4">
+        <div class="para_20">
+            <h3><a name="section09152016" id="section09152016">09/15/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Sample preparation 17: GOPTS + anti-GFP nanobody(Alexa647) + GFP</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubated with nanobody (5µl, 330µg/ml, 60 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and blocked with milk powder(5%) (30 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (10 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH2O </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubated with GFP (2µl, 1.35mg/ml, 20 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (10 min, 10rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH20</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubated with Mowiol DABCO (5µl, overnight)</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit21">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1-4</td>
+                            <td class="col1">GOPTS + nanobody(Alexa647) + GFP</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">5</td>
+                            <td class="col1">GOPTS + milk powder + GFP (negative control)</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">6</td>
+                            <td class="col1">GOPTS + milk powder + NB (negative control)</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">7</td>
+                            <td class="col1">GOPTS + GFP</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">8</td>
+                            <td class="col1">GOPTS (negative control)</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT21 TABLE [18244-18450] -->
+                <p> <strong>Fluorescence microscopy 16 (Nikon C2+ confocal microscope): GOPTS + B53</strong> </p>
+                <p> –&gt; no spores detectable </p>
+            </div>
+            <h3><a name="section09162016" id="section09162016">09/16/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Fluorescence microscopy 17 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + GFP</strong> </p>
+                <p> –&gt; GFP signal: GFP has been bound to nanobody </p>
+                <p>
+                     <center><img class="something" src="https://static.igem.org/mediawiki/2016/0/02/T--Freiburg--targetingjournal10.png"> </center>
+                   <center><img class="something" src="https://static.igem.org/mediawiki/2016/8/8c/T--Freiburg--targetingjournal11.png"> </center>
+<center><img class="something" src="https://static.igem.org/mediawiki/2016/8/83/T--Freiburg--targetingjournal20.png"> </center>
+<center><img class="something" src="https://static.igem.org/mediawiki/2016/a/a9/T--Freiburg--targetingjournal13.png"> </center>
+<center><img class="something" src="https://static.igem.org/mediawiki/2016/b/b6/T--Freiburg--targetingjournal14.png"> </center>
+<center><img class="something" src="https://static.igem.org/mediawiki/2016/2/25/T--Freiburg--targetingjournal15.png"> </center>
+<center><img class="something" src="https://static.igem.org/mediawiki/2016/e/e4/T--Freiburg--targetingjournal16.png"> </center>
+<center><img class="something" src="https://static.igem.org/mediawiki/2016/4/4b/T--Freiburg--targetingjournal17.png"> </center>
-</div>
+                    <br/> </p>
+                <ul>
+                    <li class="level1">
+                        <div class="li"> 1-4) GOPTS + nanobody(Alexa647) + GFP</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> 5) GOPTS + milk powder + GFP (negative control)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> 6) GOPTS + milk powder + NB (negative control)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> 7) GOPTS + GFP (positive control)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> 8) GOPTS (negative control)</div>
+                    </li>
+                </ul>
+                <p>
+                    <br/>
+                    <br/>
+                    <br/>
+                    <br/> </p>
+            </div>
+            <h3><a name="section09172016" id="section09172016">09/17/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Sample preparation 18: GOPTS + anti-GFP nanobody + GFP (spotting-mask)</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubation with anti-GFP nanobody ( 2µl/spot, 330µg/ml, 10 min, 4°C)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsing and blocking with milk powder(5%) (15 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsing with PBS(1x) </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubated with GFP (2µl, 1.35mg/ml, 30 min, 4°C)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (10 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH2O</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> (removed mask)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH2O</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubated with Mowiol DABCO (overnight)</div>
+                    </li>
+                </ol>
+            </div>
+        </div>
+        <!-- para_20-->
+    </div>
+    <!-- color -->
+    <div class="color5">
+        <div class="para_20">
+            <h3><a name="section09182016" id="section09182016">09/18/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Fluorescence microscopy 18 (Nikon C2+ confocal microscope): GOPTS + antiGFP-nanobody + GFP (spotting-mask)</strong> </p>
+                <p> –&gt; no GFP signal detectable </p>
+                <p> <strong>Sample preparation 19: GOPTS + anti-GFP nanobody + GFP (spotting-mask)</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubation with anti-GFP nanobody (2µl/spot, 330µg/ml, 15 min, 4°C)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsing with PBS(1x) </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> (removed mask)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsing and blocking with milk powder(5%) (15 min, 2 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> (reattached mask)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubated with GFP (2µl, 1.35mg/ml, 30 min, 4°C)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> (removed mask)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (10 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH2O</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubated with Mowiol DABCO (overnight)</div>
+                    </li>
+                </ol>
+            </div>
+            <h3><a name="section09192016" id="section09192016">09/19/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Fluorescence microscopy 19 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + GFP (spotting-mask)</strong> </p>
+                <p> –&gt; slight GFP-signal on nanobody </p>
+                <p> <strong>Sample preparation 20: anti-GFP nanobody + B53 (in solution)</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubation of anti-GFP nanobody(Alexa647) (2µl, 165µg/ml, 15 min, 4°C) + B53</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubated on GOPTS-slide (2µl, 15 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and blocked with milk powder(5%) (30 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (2 x 1 min, 10 rpm)</div>
+                    </li>
+                </ol>
+                <p> –&gt; problem: no washing away of unbound nanobody </p>
+                <div class="table sectionedit22">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1,2</td>
+                            <td class="col1">B53 + nanobody(Alexa647) (incubated in solution) on GOPTS</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">3,4</td>
+                            <td class="col1">WT + nanobody(Alexa647) (incubated in solution) on GOPTS (negative control)</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">5,6</td>
+                            <td class="col1">B53 + nanobody(Alexa647) (incubated in solution) dried on milk powder</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">7,8</td>
+                            <td class="col1">WT + nanobody(Alexa647) (incubated in solution) dried on milk powder (negative control)</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">9,10</td>
+                            <td class="col1">GOPTS + nanobody(Alexa647) + GFP (positive control)</td>
+                        </tr>
+                        <tr class="row6">
+                            <td class="col0">11,12</td>
+                            <td class="col1">GOPTS + nanobody(Alexa647) + B53</td>
+                        </tr>
+                        <tr class="row7">
+                            <td class="col0">13,14</td>
+                            <td class="col1">GOPTS + nanobody(Alexa647) + WT (negative control)</td>
+                        </tr>
+                        <tr class="row8">
+                            <td class="col0">15</td>
+                            <td class="col1">GOPTS + milk powder + nanobody(Alexa647) (negative control)</td>
+                        </tr>
+                        <tr class="row9">
+                            <td class="col0">16</td>
+                            <td class="col1">GOPTS + milk powder + B53 (negative control)</td>
+                        </tr>
+                        <tr class="row10">
+                            <td class="col0">17</td>
+                            <td class="col1">GOPTS + milk powder + WT (negative control)</td>
+                        </tr>
+                        <tr class="row11">
+                            <td class="col0">18</td>
+                            <td class="col1">GOPTS + GFP (positive control)</td>
+                        </tr>
+                        <tr class="row12">
+                            <td class="col0">19</td>
+                            <td class="col1">GOPTS + nanobody(Alexa647) (negative control)</td>
+                        </tr>
+                        <tr class="row13">
+                            <td class="col0">20</td>
+                            <td class="col1">B53 dried (positive control)</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT22 TABLE [21203-21988] -->
+            </div>
+            <h3><a name="section09202016" id="section09202016">09/20/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Sample preparation 21: anti-GFP nanobody + B53 (in solution)</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubation with anti-GFP nanobody(Alexa647) (2µl, 165µg/ml, 30 min, 4°C) + B53</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> centrifuged (2 min, 13400 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> washed with PBS(1x)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> repeated steps 2 and 3 </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> resuspended in PBS(1x) (30µl)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubated on GOPTS-slide (2µl, 15 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and blocked with milkpowder(5%) (30 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (2 x 1 min, 10 rpm)</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit23">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1</td>
+                            <td class="col1">B53 + nanobody(Alexa647) (incubated in solution) on GOPTS</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">2</td>
+                            <td class="col1">WT + nanobody(Alexa647) (incubated in solution) on GOPTS</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">3,4</td>
+                            <td class="col1">B53 + nanobody(Alexa647) dried on milkpowder</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">5,6</td>
+                            <td class="col1">WT + nanobody(Alexa647) dried on milkpowder</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT23 TABLE [22443-22692] -->
+            </div>
+        </div>
+        <!-- para_20-->
+    </div>
+    <!-- color -->
+    <div class="color6">
+        <div class="para_20">
+            <h3><a name="section09292016" id="section09292016">09/29/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Sample preparation 22: GOPTS + GFP</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubation with GFP (1 µl, 1.35mg/ml, 30 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and blocked with milk powder(5%) (30 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (10 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH2O </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with Mowiol DABCO (5µl, overnight)</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit24">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0 leftalign">1-5 </td>
+                            <td class="col1">GOPTS + GFP</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0 leftalign">6 </td>
+                            <td class="col1">GOPTS</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT24 TABLE [23013-23068] -->
+            </div>
+            <h3><a name="section09302016" id="section09302016">09/30/2016</a></h3>
+            <div class="level4">
+                <p> <em class="u"><strong>–&gt; Construct spores are ready for analysis</strong></em> </p>
+                <div class="table sectionedit25">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Strain No.</strong></td>
+                            <td class="col1"><strong>Strain</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">3</td>
+                            <td class="col1">150: aGFPnano_HA_aHelix_cgeA</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">6</td>
+                            <td class="col1">151: aGFPnano_HA_aHelix_cotG</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">7</td>
+                            <td class="col1">159: aGFPnano_HA_G4S_CotZ</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">9</td>
+                            <td class="col1">157: aGFPnano_HA_aHelix_CotZ</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT25 TABLE [23141-23297] -->
+                <p> <strong>Fluorescence microscopy 22 (Nikon C2+ confocal microscope): GOPTS + GFP</strong> </p>
+                <p> –&gt; spots with evenly distributed GFP </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubation with GFP (1 µl, 1.35mg/ml, 30 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> spores centrifuged (2 min, 13,400 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> washed with PBS(1x)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> step 1 repeated</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> resuspended in PBS(1x)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation on GOPTS-slide(GFP) with spores (4µl, 400,000/µl, incubation time: see table below, 4°C)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and blocked with milk powder(5%) (30 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with PBS(1x) </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH2O</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with Mowiol DABCO (overnight)</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit26">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                            <td class="col2"><strong>Incubation time (min)</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1</td>
+                            <td class="col1">GOPTS + GFP + strain 3</td>
+                            <td class="col2">5</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">2</td>
+                            <td class="col1">GOPTS + GFP + strain 3</td>
+                            <td class="col2">10</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">3</td>
+                            <td class="col1">GOPTS + GFP + strain 3</td>
+                            <td class="col2">15</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">4</td>
+                            <td class="col1">GOPTS + GFP + strain 6</td>
+                            <td class="col2">5</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">5</td>
+                            <td class="col1">GOPTS + GFP + strain 6</td>
+                            <td class="col2">10</td>
+                        </tr>
+                        <tr class="row6">
+                            <td class="col0">6</td>
+                            <td class="col1">GOPTS + GFP + strain 6</td>
+                            <td class="col2">15</td>
+                        </tr>
+                        <tr class="row7">
+                            <td class="col0">7</td>
+                            <td class="col1">GOPTS + GFP + strain 7</td>
+                            <td class="col2">5</td>
+                        </tr>
+                        <tr class="row8">
+                            <td class="col0">8</td>
+                            <td class="col1">GOPTS + GFP + strain 7</td>
+                            <td class="col2">10</td>
+                        </tr>
+                        <tr class="row9">
+                            <td class="col0">9</td>
+                            <td class="col1">GOPTS + GFP + strain 7</td>
+                            <td class="col2">15</td>
+                        </tr>
+                        <tr class="row10">
+                            <td class="col0">10</td>
+                            <td class="col1">GOPTS + GFP + strain 9</td>
+                            <td class="col2">5</td>
+                        </tr>
+                        <tr class="row11">
+                            <td class="col0">11</td>
+                            <td class="col1">GOPTS + GFP + strain 9</td>
+                            <td class="col2">10</td>
+                        </tr>
+                        <tr class="row12">
+                            <td class="col0">12</td>
+                            <td class="col1">GOPTS + GFP + strain 9</td>
+                            <td class="col2">15</td>
+                        </tr>
+                        <tr class="row13">
+                            <td class="col0">13, 14</td>
+                            <td class="col1">GOPTS + GFP (positive control)</td>
+                            <td class="col2">10</td>
+                        </tr>
+                        <tr class="row14">
+                            <td class="col0">15</td>
+                            <td class="col1">GOPTS + GFP + WT (negative control)</td>
+                            <td class="col2">5</td>
+                        </tr>
+                        <tr class="row15">
+                            <td class="col0">16</td>
+                            <td class="col1">GOPTS + GFP + WT (negative control)</td>
+                            <td class="col2">10</td>
+                        </tr>
+                        <tr class="row16">
+                            <td class="col0">17</td>
+                            <td class="col1">GOPTS + GFP + WT (negative control)</td>
+                            <td class="col2">15</td>
+                        </tr>
+                        <tr class="row17">
+                            <td class="col0">18</td>
+                            <td class="col1">GOPTS + strain 3 (positive control)</td>
+                            <td class="col2">10</td>
+                        </tr>
+                        <tr class="row18">
+                            <td class="col0">19</td>
+                            <td class="col1">GOPTS + strain 6 (positive control)</td>
+                            <td class="col2">10</td>
+                        </tr>
+                        <tr class="row19">
+                            <td class="col0">20</td>
+                            <td class="col1">GOPTS + strain 7 (positive control)</td>
+                            <td class="col2">10</td>
+                        </tr>
+                        <tr class="row20">
+                            <td class="col0">21</td>
+                            <td class="col1">GOPTS + strain 9 (positive control)</td>
+                            <td class="col2">10</td>
+                        </tr>
+                        <tr class="row21">
+                            <td class="col0">22</td>
+                            <td class="col1">GOPTS + WT (negative control)</td>
+                            <td class="col2">10</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT26 TABLE [23872-24670] -->
+            </div>
+        </div>
+        <!-- para_20-->
+    </div>
+    <!-- color -->
+    <div class="color7">
+        <div class="para_20">
+            <h3><a name="section10012016" id="section10012016">10/01/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Sample preparation 23: GOPTS + GFP + spores</strong> </p>
+                <p> –&gt; no spores detectable </p>
+                <p> <strong>Sample preparation 24: GOPTS + GFP + spores (in EtOH resuspended)</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubation of GFP (4µl, 1,35g/ml) + XXX (4µl, 400,000/µl) (10 min, 4°C)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> solution centrifuged (4 min, 13,400 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> washed with PBS(1x)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> step 2 repeated</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> resuspended in EtOH (250µl)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation on GOPTS-slide (4µl, 30 min, 4°C)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and blocked with milk powder(5%) (30 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with PBS(1x) </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH2O</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with Mowiol DABCO (overnight)</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit27">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1-3</td>
+                            <td class="col1">GOPTS + GFP + strain 6</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">4-6</td>
+                            <td class="col1">GOPTS + GFP + WT</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">7-9</td>
+                            <td class="col1">GOPTS + GFP</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">10-12</td>
+                            <td class="col1">GOPTS + strain 6</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">13-15</td>
+                            <td class="col1">GOPTS + WT</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT27 TABLE [25275-25414] -->
+            </div>
+            <h3><a name="section10022016" id="section10022016">10/02/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Fluorescence microscopy 24 (Nikon C2+ confocal microscope): GOPTS + GFP + spores (strains 3,6,7,9,wt) (in EtOH resuspended)</strong> </p>
+                <p> –&gt; EtOH denaturated GFP: very weak signal. No spores detectable </p>
+                <p> <strong>Sample preparation 25: GFP + spores (strains 3,6,7,9) (in solution)</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubation of GFP (4µl, 1,35g/ml) + spores(strain 3,6,7,9) (4µl, 400,000/µl) (incubation time: see table below, 4°C)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> centrifuged (2 min, 13400 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> washed with PBS(1x)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> repeated steps 2 and 3 </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> resuspended in PBS(1x) (20µl)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubated on GOPTS-slide (4µl, 10 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and blocked with milk powder(5%) (30 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (10 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH2O</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubrnight)ation with Mowiol DABCO </div>
+                    </li>
+                </ol>
+                <div class="table sectionedit28">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1, 2</td>
+                            <td class="col1">GFP + strain 3</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">3, 4</td>
+                            <td class="col1">GFP + strain 6</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">5, 6</td>
+                            <td class="col1">GFP + strain 7</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">7, 8</td>
+                            <td class="col1">GFP + strain 9</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">9, 10</td>
+                            <td class="col1">GFP + WT (negative control)</td>
+                        </tr>
+                        <tr class="row6">
+                            <td class="col0">11, 12</td>
+                            <td class="col1">GFP + PBS (positive control)</td>
+                        </tr>
+                        <tr class="row7">
+                            <td class="col0">13, 14</td>
+                            <td class="col1">GFP (positive control)</td>
+                        </tr>
+                        <tr class="row8">
+                            <td class="col0">15, 16</td>
+                            <td class="col1">WT (negative control)</td>
+                        </tr>
+                        <tr class="row9">
+                            <td class="col0">17</td>
+                            <td class="col1">strain 3 (negative control)</td>
+                        </tr>
+                        <tr class="row10">
+                            <td class="col0">18</td>
+                            <td class="col1">strain 6 (negative control)</td>
+                        </tr>
+                        <tr class="row11">
+                            <td class="col0">19</td>
+                            <td class="col1">strain 7 (negative control)</td>
+                        </tr>
+                        <tr class="row12">
+                            <td class="col0">20</td>
+                            <td class="col1">strain 9 (negative control)</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT28 TABLE [26200-26582] -->
+            </div>
+            <h3><a name="section10032016" id="section10032016">10/03/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Sample preparation 26: Incubation test: GFP + spores (in solution)</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubation of GFP (4µl, 1,35g/ml) + spores (4µl, 400,000/µl) (incubation time: see tables below, 4°C)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> centrifuged (2 min, 13400 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> washed with PBS(1x)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> repeated steps 2 and 3)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> resuspended in PBS(1x) (20µl))</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubated on GOPTS-slide (4µl, 10 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and blocked with milk powder(5%) (30 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (10 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH2O</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with Mowiol DABCO (overnight)</div>
+                    </li>
+                </ol>
+                <p> Slide 1: </p>
+                <div class="table sectionedit29">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                            <td class="col2"><strong>Incubation time</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1, 2</td>
+                            <td class="col1">GFP + strain 3</td>
+                            <td class="col2">30 min</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">3, 4</td>
+                            <td class="col1">GFP + strain 3(pellet washed with PBS(1x))</td>
+                            <td class="col2">30 min</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">5, 6</td>
+                            <td class="col1">GFP + strain 6</td>
+                            <td class="col2">30 min</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">7, 8</td>
+                            <td class="col1">GFP + strain 6(pellet washed with PBS(1x))</td>
+                            <td class="col2">30 min</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">9, 10</td>
+                            <td class="col1">GFP + strain 7</td>
+                            <td class="col2">30 min</td>
+                        </tr>
+                        <tr class="row6">
+                            <td class="col0">11, 12</td>
+                            <td class="col1">GFP + strain 7(pellet washed with PBS(1x))</td>
+                            <td class="col2">30 min</td>
+                        </tr>
+                        <tr class="row7">
+                            <td class="col0">13, 14</td>
+                            <td class="col1">GFP + strain 9</td>
+                            <td class="col2">30 min</td>
+                        </tr>
+                        <tr class="row8">
+                            <td class="col0">15, 16</td>
+                            <td class="col1">GFP + strain 9 (pellet washed withn PBS(1x))</td>
+                            <td class="col2">30 min</td>
+                        </tr>
+                        <tr class="row9">
+                            <td class="col0">17, 18</td>
+                            <td class="col1">GFP + WT (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row10">
+                            <td class="col0">19, 20</td>
+                            <td class="col1">GFP + WT (negative control) (pellet washed with PBS(1x))</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row11">
+                            <td class="col0">21, 22</td>
+                            <td class="col1">GFP (positive control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row12">
+                            <td class="col0">23, 24</td>
+                            <td class="col1">WT (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row13">
+                            <td class="col0">25</td>
+                            <td class="col1">strain 3 (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row14">
+                            <td class="col0">26</td>
+                            <td class="col1">strain 6 (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row15">
+                            <td class="col0">27</td>
+                            <td class="col1">strain 7 (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row16">
+                            <td class="col0">28</td>
+                            <td class="col1">strain 9 (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT29 TABLE [27172-27869] -->
+                <p> Slide 2: </p>
+                <div class="table sectionedit30">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                            <td class="col2"><strong>Incubation time</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1, 2</td>
+                            <td class="col1">GFP + strain 3</td>
+                            <td class="col2">60 min</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">3, 4</td>
+                            <td class="col1">GFP + strain 3(pellet washed with PBS(1x))</td>
+                            <td class="col2">60 min</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">5, 6</td>
+                            <td class="col1">GFP + strain 6</td>
+                            <td class="col2">60 min</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">7, 8</td>
+                            <td class="col1">GFP + strain 6(pellet washed with PBS(1x))</td>
+                            <td class="col2">60 min</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">9, 10</td>
+                            <td class="col1">GFP + strain 7</td>
+                            <td class="col2">60 min</td>
+                        </tr>
+                        <tr class="row6">
+                            <td class="col0">11, 12</td>
+                            <td class="col1">GFP + strain 7(pellet washed with PBS(1x))</td>
+                            <td class="col2">60 min</td>
+                        </tr>
+                        <tr class="row7">
+                            <td class="col0">13, 14</td>
+                            <td class="col1">GFP + strain 9</td>
+                            <td class="col2">60 min</td>
+                        </tr>
+                        <tr class="row8">
+                            <td class="col0">15, 16</td>
+                            <td class="col1">GFP + strain 9 (pellet washed withn PBS(1x))</td>
+                            <td class="col2">60 min</td>
+                        </tr>
+                        <tr class="row9">
+                            <td class="col0">17, 18</td>
+                            <td class="col1">GFP + WT (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row10">
+                            <td class="col0">19, 20</td>
+                            <td class="col1">GFP + WT (negative control) (pellet washed with PBS(1x))</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row11">
+                            <td class="col0">21, 22</td>
+                            <td class="col1">GFP (positive control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row12">
+                            <td class="col0">23, 24</td>
+                            <td class="col1">WT (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row13">
+                            <td class="col0">25</td>
+                            <td class="col1">strain 3 (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row14">
+                            <td class="col0">26</td>
+                            <td class="col1">strain 6 (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row15">
+                            <td class="col0">27</td>
+                            <td class="col1">strain 7 (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row16">
+                            <td class="col0">28</td>
+                            <td class="col1">strain 9 (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT30 TABLE [27881-28578] -->
+                <p> Slide 3: </p>
+                <div class="table sectionedit31">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                            <td class="col2"><strong>Incubation time</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1, 2</td>
+                            <td class="col1">GFP + strain 3</td>
+                            <td class="col2">120 min</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">3, 4</td>
+                            <td class="col1">GFP + strain 3(pellet washed with PBS(1x))</td>
+                            <td class="col2">120 min</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">5, 6</td>
+                            <td class="col1">GFP + strain 6</td>
+                            <td class="col2">120 min</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">7, 8</td>
+                            <td class="col1">GFP + strain 6(pellet washed with PBS(1x))</td>
+                            <td class="col2">120 min</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">9, 10</td>
+                            <td class="col1">GFP + strain 7</td>
+                            <td class="col2">120 min</td>
+                        </tr>
+                        <tr class="row6">
+                            <td class="col0">11, 12</td>
+                            <td class="col1">GFP + strain 7(pellet washed with PBS(1x))</td>
+                            <td class="col2">120 min</td>
+                        </tr>
+                        <tr class="row7">
+                            <td class="col0">13, 14</td>
+                            <td class="col1">GFP + strain 9</td>
+                            <td class="col2">120 min</td>
+                        </tr>
+                        <tr class="row8">
+                            <td class="col0">15, 16</td>
+                            <td class="col1">GFP + strain 9 (pellet washed withn PBS(1x))</td>
+                            <td class="col2">120 min</td>
+                        </tr>
+                        <tr class="row9">
+                            <td class="col0">17, 18</td>
+                            <td class="col1">GFP + WT (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row10">
+                            <td class="col0">19, 20</td>
+                            <td class="col1">GFP + WT (negative control) (pellet washed with PBS(1x))</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row11">
+                            <td class="col0">21, 22</td>
+                            <td class="col1">GFP (positive control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row12">
+                            <td class="col0">23, 24</td>
+                            <td class="col1">WT (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row13">
+                            <td class="col0">25</td>
+                            <td class="col1">strain 3 (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row14">
+                            <td class="col0">26</td>
+                            <td class="col1">strain 6 (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row15">
+                            <td class="col0">27</td>
+                            <td class="col1">strain 7 (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row16">
+                            <td class="col0">28</td>
+                            <td class="col1">strain 9 (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT31 TABLE [28590-29295] -->
+                <p> <strong>Sample preparation 28: GFP-volume test B: GFP + strain 7 (in solution, washed)</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubation of GFP (volume: see table below, 1,35g/ml) + strain 7 (4µl, 400,000/µl) (90 min, 4°C)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> centrifuged (2 min, 13400 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> washed with PBS(1x)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> repeated steps 2 and 3)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> resuspended in PBS(1x) (20µl)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubated on GOPTS-slide (2µl, 90 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and blocked with milk powder(5%) (30 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (10 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH2O</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with Mowiol DABCO (overnight)</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit32">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                            <td class="col2"><strong>Volume (GFP)</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1, 2</td>
+                            <td class="col1">GFP + strain 7</td>
+                            <td class="col2">0,5µl</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">3, 4</td>
+                            <td class="col1">GFP + strain 7</td>
+                            <td class="col2">1,0µl</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">5, 6</td>
+                            <td class="col1">GFP + strain 7</td>
+                            <td class="col2">1,5µl</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">7, 8</td>
+                            <td class="col1">GFP + strain 7</td>
+                            <td class="col2">2,0µl</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">9, 10</td>
+                            <td class="col1">GFP + strain 7</td>
+                            <td class="col2">2,5µl</td>
+                        </tr>
+                        <tr class="row6">
+                            <td class="col0">11, 12</td>
+                            <td class="col1">GFP (positive control)</td>
+                            <td class="col2">4,0µl</td>
+                        </tr>
+                        <tr class="row7">
+                            <td class="col0">13, 14</td>
+                            <td class="col1">strain 7 (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row8">
+                            <td class="col0">15, 16</td>
+                            <td class="col1">GFP + WT (negative control)</td>
+                            <td class="col2">0,5µl</td>
+                        </tr>
+                        <tr class="row9">
+                            <td class="col0">17, 18</td>
+                            <td class="col1">GFP + WT (negative control)</td>
+                            <td class="col2">1,0µl</td>
+                        </tr>
+                        <tr class="row10">
+                            <td class="col0">19, 20</td>
+                            <td class="col1">GFP + WT (negative control)</td>
+                            <td class="col2">1,5µl</td>
+                        </tr>
+                        <tr class="row11">
+                            <td class="col0">21, 22</td>
+                            <td class="col1">GFP + WT (negative control)</td>
+                            <td class="col2">2,0µl</td>
+                        </tr>
+                        <tr class="row12">
+                            <td class="col0">23, 24</td>
+                            <td class="col1">GFP + WT (negative control)</td>
+                            <td class="col2">2,5µl</td>
+                        </tr>
+                        <tr class="row13">
+                            <td class="col0">25, 26</td>
+                            <td class="col1">WT (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT32 TABLE [29862-30378] -->
+                <p> <strong>Sample preparation 29: Spore-amount test: GFP + strain 7 (in solution)</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubation of GFP (0,5µl, 1,35g/ml) + strain 7 (amount: see table below, 400,000/µl) (90 min, 4°C)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> centrifuged (2 min, 13400 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> washed with PBS(1x)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> repeated steps 2 and 3)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> resuspended in PBS(1x) (20µl)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubated on GOPTS-slide (2µl, 90 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and blocked with milk powder(5%) (30 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and washed with PBS(1x) (10 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH2O</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with Mowiol DABCO (overnight)</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit33">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                            <td class="col2"><strong>Amount (spores)</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1, 2</td>
+                            <td class="col1">strain 7 (negative control)</td>
+                            <td class="col2">25,000,000</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">3, 4</td>
+                            <td class="col1">strain 7 (negative control)</td>
+                            <td class="col2">15,000,000</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">5, 6</td>
+                            <td class="col1">strain 7 (negative control)</td>
+                            <td class="col2">10,000,000</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">7, 8</td>
+                            <td class="col1">GFP + strain 7</td>
+                            <td class="col2">25,000,000</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">9, 10</td>
+                            <td class="col1">GFP + strain 7</td>
+                            <td class="col2">15,000,000</td>
+                        </tr>
+                        <tr class="row6">
+                            <td class="col0">11, 12</td>
+                            <td class="col1">GFP + strain 7</td>
+                            <td class="col2">10,000,000</td>
+                        </tr>
+                        <tr class="row7">
+                            <td class="col0">13, 14</td>
+                            <td class="col1">GFP (positive control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row8">
+                            <td class="col0">15, 16</td>
+                            <td class="col1">WT (negative control)</td>
+                            <td class="col2">25,000,000</td>
+                        </tr>
+                        <tr class="row9">
+                            <td class="col0">17, 18</td>
+                            <td class="col1">WT (negative control)</td>
+                            <td class="col2">15,000,000</td>
+                        </tr>
+                        <tr class="row10">
+                            <td class="col0">19, 20</td>
+                            <td class="col1">WT(negative control)</td>
+                            <td class="col2">10,000,000</td>
+                        </tr>
+                        <tr class="row11">
+                            <td class="col0">21, 22</td>
+                            <td class="col1">GFP + WT (negative control)</td>
+                            <td class="col2">25,000,000</td>
+                        </tr>
+                        <tr class="row12">
+                            <td class="col0">23, 24</td>
+                            <td class="col1">GFP + WT (negative control)</td>
+                            <td class="col2">15,000,000</td>
+                        </tr>
+                        <tr class="row13">
+                            <td class="col0">25, 26</td>
+                            <td class="col1">GFP + WT (negative control)</td>
+                            <td class="col2">10,000,000</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT33 TABLE [30939-31526] -->
+            </div>
+        </div>
+        <!-- para_20-->
+    </div>
+    <!-- color -->
+    <div class="color8">
+        <div class="para_20">
+            <h3><a name="section10042016" id="section10042016">10/04/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Fluorescence microscopy 24 (Nikon C2+ confocal microscope): GOPTS + GFP + spores (in EtOH resuspended)</strong> </p>
+                <p> –&gt; spore concentration too high, too many layers of spores </p>
+                <p> <strong>Fluorescence microscopy 25 (Nikon C2+ confocal microscope): GFP + spores (in solution)</strong> </p>
+                <p> –&gt; spore concentration too high, too many layers of spores </p>
+                <p> <strong>Fluorescence microscopy 26 (Nikon C2+ confocal microscope): Incubation test: GFP + spores (in solution)</strong> </p>
+                <p> –&gt; spore concentration too high, too many layers of spores </p>
+                <p> Slide 1: </p>
+                <p>
+                    <center><img class="something" src="https://static.igem.org/mediawiki/2016/f/f5/T--Freiburg--targetingjournal18.png"> </center>
+<center><img class="something" src="https://static.igem.org/mediawiki/2016/a/a3/T--Freiburg--targetingjournal19.png"> </center>
+<center><img class="something" src="https://static.igem.org/mediawiki/2016/9/90/T--Freiburg--targetingjournal12.png"> </center>
+<center><img class="something" src="https://static.igem.org/mediawiki/2016/9/9f/T--Freiburg--targetingjournal21.png"> </center>
-</div>
+                    <br/> </p>
+                <p> 1.1) strain7 + GFP (30 min) (white field) 1.2) strain7 + GFP (30 min) (GFP-channel) 2.1) strain9 + GFP (30 min) (white field) 2.2) strain9 + GFP (30 min) (GFP-channel)
+                    <br/>
+                    <br/>
+                    <br/>
+                    <br/>
+                    <br/>
+                    <br/>
+                    <br/>
+                    <br/> Slide 2:
+                    <br/>
+                   <center><img class="something" src="https://static.igem.org/mediawiki/2016/6/61/T--Freiburg--targetingjournal22.png"> </center>
+                   <center><img class="something" src="https://static.igem.org/mediawiki/2016/b/be/T--Freiburg--targetingjournal23.png"> </center>
+                   <center><img class="something" src="https://static.igem.org/mediawiki/2016/8/88/T--Freiburg--targetingjournal24.png"> </center>
+                   <center><img class="something" src="https://static.igem.org/mediawiki/2016/1/13/T--Freiburg--targetingjournal25.png"> </center>
-</div>
+                    <br/> </p>
+                <p> 1.1) strain 7 + GFP (90 min)(white field) 1.2) strain 7 + GFP (90 min)(GFP-channel) 2.1) strain WT + GFP (90 min)(white field) 2.2) strain WT + GFP (90 min)(GFP-channel)
+                    <br/>
+                    <br/>
+                    <br/>
+                    <br/>
+                    <br/>
+                    <br/>
+                    <br/>
+                    <br/> <strong>Fluorescence microscopy 27 (Nikon C2+ confocal microscope): GFP-volume test A: GFP + strain 7 (in solution)</strong> </p>
+                <p> –&gt; background fluorescence was too high </p>
+                <p> <strong>Fluorescence microscopy 28 (Nikon C2+ confocal microscope): GFP-volume test B: GFP + strain 7 (in solution)</strong> </p>
+                <p> –&gt; background fluorescence was too high </p>
+                <p> <strong>Fluorescence microscopy 29 (Nikon C2+ confocal microscope): GFP + strain 7 (in solution)</strong> </p>
+                <p> –&gt; spores and WT show similar levels of fluorescence, probably autofluorescence </p>
+                <p>
+                   <center><img class="something" src="https://static.igem.org/mediawiki/2016/b/ba/T--Freiburg--targetingjournal26.png"> </center>
+                   <center><img class="something" src="https://static.igem.org/mediawiki/2016/8/83/T--Freiburg--targetingjournal27.png"> </center>
+                   <center><img class="something" src="https://static.igem.org/mediawiki/2016/8/88/T--Freiburg--targetingjournal28.png"> </center>
+                   <center><img class="something" src="https://static.igem.org/mediawiki/2016/1/1b/T--Freiburg--targetingjournal29.png"> </center>
-</div>
+                    <br/> </p>
+                <p> 1) strain 7 + GFP (white field) 2) strain 7 + GFP (GFP-channel) 3) WT + GFP (white field) 4) WT + GFP (GFP-channel)
+                    <br/>
+                    <br/>
+                    <br/>
+                    <br/>
+                    <br/>
+                    <br/>
+                    <br/>
+                    <br/> <strong>Sample preparation 30: Wash-test A: GFP + spores (strains 3,6,7,9) (in solution)</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubation of GFP (1µl, 1,35g/ml) + spores (strain 3,6,7,9) (25,000.000 spores) (90 min, 4°C)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> solution centrifuged (2 min, 13,400 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> washed with PBS(1x)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> step 2 and 3 repeated (2x/4x)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> resuspended in EtOH (250µl)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation on GOPTS-slide (4µl, 30 min, 4°C)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and blocked with milk powder(5%) (30 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with PBS(1x) </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH2O</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with Mowiol DABCO (overnight)</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit34">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                            <td class="col2"><strong>Washing of GFP-spore solution</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1, 2</td>
+                            <td class="col1">GFP + strain 3</td>
+                            <td class="col2">2x</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">3, 4</td>
+                            <td class="col1">GFP + strain 6</td>
+                            <td class="col2">2x</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">5, 6</td>
+                            <td class="col1">GFP + strain 7</td>
+                            <td class="col2">2x</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">7, 8</td>
+                            <td class="col1">GFP + strain 9</td>
+                            <td class="col2">2x</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">9, 10</td>
+                            <td class="col1">GFP + WT (negative control)</td>
+                            <td class="col2">2x</td>
+                        </tr>
+                        <tr class="row6">
+                            <td class="col0">11, 12</td>
+                            <td class="col1">GFP + strain 3</td>
+                            <td class="col2">4x</td>
+                        </tr>
+                        <tr class="row7">
+                            <td class="col0">13, 14</td>
+                            <td class="col1">GFP + strain 6</td>
+                            <td class="col2">4x</td>
+                        </tr>
+                        <tr class="row8">
+                            <td class="col0">15, 16</td>
+                            <td class="col1">GFP + strain 7</td>
+                            <td class="col2">4x</td>
+                        </tr>
+                        <tr class="row9">
+                            <td class="col0">17, 18</td>
+                            <td class="col1">GFP + strain 9</td>
+                            <td class="col2">4x</td>
+                        </tr>
+                        <tr class="row10">
+                            <td class="col0">19, 20</td>
+                            <td class="col1">GFP + WT (negative control)</td>
+                            <td class="col2">4x</td>
+                        </tr>
+                        <tr class="row11">
+                            <td class="col0">21, 22</td>
+                            <td class="col1">GFP (positive control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row12">
+                            <td class="col0">23</td>
+                            <td class="col1">WT (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row13">
+                            <td class="col0">24</td>
+                            <td class="col1">strain 3 (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row14">
+                            <td class="col0">25</td>
+                            <td class="col1">strain 6 (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row15">
+                            <td class="col0">26</td>
+                            <td class="col1">strain 7 (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row16">
+                            <td class="col0">27</td>
+                            <td class="col1">strain 9 (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT34 TABLE [34545-35086] -->
+                <p> <strong>Sample preparation 31: Wash-test B: GFP + spores (in solution)</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubation of GFP (1µl, 1,35g/ml) + spores (25,000.000 spores) (90 min, 4°C)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> solution centrifuged (2 min, 13,400 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> washed with TBST(0,005%)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> steps 2 and 3 repeated (2x/4x)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> resuspended in EtOH (250µl)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation on GOPTS-slide (4µl, 30 min, 4°C)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and blocked with milk powder(5%) (30 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with PBS(1x) </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH2O</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with Mowiol DABCO (overnight)</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit35">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                            <td class="col2"><strong>Washing of GFP-spore solution</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1, 2</td>
+                            <td class="col1">GFP + strain 3</td>
+                            <td class="col2">2x</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">3, 4</td>
+                            <td class="col1">GFP + strain 6</td>
+                            <td class="col2">2x</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">5, 6</td>
+                            <td class="col1">GFP + strain 7</td>
+                            <td class="col2">2x</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">7, 8</td>
+                            <td class="col1">GFP + strain 9</td>
+                            <td class="col2">2x</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">9, 10</td>
+                            <td class="col1">GFP + WT (negative control)</td>
+                            <td class="col2">2x</td>
+                        </tr>
+                        <tr class="row6">
+                            <td class="col0">11, 12</td>
+                            <td class="col1">GFP + strain 3</td>
+                            <td class="col2">4x</td>
+                        </tr>
+                        <tr class="row7">
+                            <td class="col0">13, 14</td>
+                            <td class="col1">GFP + strain 6</td>
+                            <td class="col2">4x</td>
+                        </tr>
+                        <tr class="row8">
+                            <td class="col0">15, 16</td>
+                            <td class="col1">GFP + strain 7</td>
+                            <td class="col2">4x</td>
+                        </tr>
+                        <tr class="row9">
+                            <td class="col0">17, 18</td>
+                            <td class="col1">GFP + strain 9</td>
+                            <td class="col2">4x</td>
+                        </tr>
+                        <tr class="row10">
+                            <td class="col0">19, 20</td>
+                            <td class="col1">GFP + WT (negative control)</td>
+                            <td class="col2">4x</td>
+                        </tr>
+                        <tr class="row11">
+                            <td class="col0">21, 22</td>
+                            <td class="col1">GFP (positive control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row12">
+                            <td class="col0">23</td>
+                            <td class="col1">WT (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row13">
+                            <td class="col0">24</td>
+                            <td class="col1">strain 3 (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row14">
+                            <td class="col0">25</td>
+                            <td class="col1">strain 6 (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row15">
+                            <td class="col0">26</td>
+                            <td class="col1">strain 7 (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row16">
+                            <td class="col0">27</td>
+                            <td class="col1">strain 9 (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT35 TABLE [35617-36158] -->
+            </div>
+        </div>
+        <!-- para_20-->
+    </div>
+    <!-- color -->
+    <div class="color1">
+        <div class="para_20">
+            <h3><a name="section10052016" id="section10052016">10/05/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Fluorescence microscopy 30 (Nikon C2+ confocal microscope): Wash-test A: GFP + spores (strains 3,6,7,9) (in solution)</strong> </p>
+                <p> –&gt; spore concentration too high, background fluorescence too high </p>
+                <p> <strong>Fluorescence microscopy 31 (Nikon C2+ confocal microscope): Wash-test B: GFP + spores (strains 3,6,7,9) (in solution)</strong> </p>
+                <p> –&gt; indicates that strain 3 binds GFP </p>
+                <p>
+                    <a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:1005.st3.gfp.slide27.waschtest4x.100ms60xj.jpg" class="media" title="labprotocol:1005.st3.gfp.slide27.waschtest4x.100ms60xj.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:1005.st3.gfp.slide27.waschtest4x.100ms60xj.jpg" class="medialeft" align="left" alt="" width="200" /></a>
+                    <a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:1005.st3.gfp.slide27.waschtest4x.4s60xj.jpg" class="media" title="labprotocol:1005.st3.gfp.slide27.waschtest4x.4s60xj.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:1005.st3.gfp.slide27.waschtest4x.4s60xj.jpg" class="medialeft" align="left" alt="" width="200" /></a>
+                    <a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:1005.wt.gfp.slide27.waschtest4x.100ms60xd.jpg" class="media" title="labprotocol:1005.wt.gfp.slide27.waschtest4x.100ms60xd.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:1005.wt.gfp.slide27.waschtest4x.100ms60xd.jpg" class="medialeft" align="left" alt="" width="200" /></a>
+                    <a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:1005.wt.gfp.slide27.waschtest4x.4s60xd.jpg" class="media" title="labprotocol:1005.wt.gfp.slide27.waschtest4x.4s60xd.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:1005.wt.gfp.slide27.waschtest4x.4s60xd.jpg" class="medialeft" align="left" alt="" width="200" /></a>
+                    <a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:1005.slide_27-wt_ohne_gfp-kontrolle.slide27-.4s60xi.jpg" class="media" title="labprotocol:1005.slide_27-wt_ohne_gfp-kontrolle.slide27-.4s60xi.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:1005.slide_27-wt_ohne_gfp-kontrolle.slide27-.4s60xi.jpg" class="medialeft" align="left" alt="" width="200" /></a>
+                    <br/> </p>
+                <ul>
+                    <li class="level1">
+                        <div class="li"> 1.1) GFP + strain 3 (white field)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> 1.2) GFP + strain 3 (GFP-channel)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> 2.1) GFP + WT (negative control) (white field)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> 2.2) GFP + WT (negative control) (GFP-channel)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> 3) WT (negative control)</div>
+                    </li>
+                </ul>
+                <p>
+                    <br/> </p>
+            </div>
+            <h3><a name="section10062016" id="section10062016">10/06/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Sample preparation 32: Wash-test C: GFP + spores (in solution)</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> incubation of GFP (1µl, 1,35g/ml) + spores (25,000.000 spores) (90 min, 4°C)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> solution centrifuged (2 min, 13,400 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> washed with TBST(0,005%)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> steps 2 and 3 repeated (1x/3x/5x)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> resuspended in EtOH (250µl)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation on GOPTS-slide (4µl, 30 min, 4°C)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and blocked with milk powder(5%) (30 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with PBS(1x) </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH2O</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with Mowiol DABCO (overnight)</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit36">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                            <td class="col2"><strong>Washing of GFP-spore solution (step 3)</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1, 2</td>
+                            <td class="col1">GFP + strain 3</td>
+                            <td class="col2">1x</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">3, 4</td>
+                            <td class="col1">GFP + strain 7</td>
+                            <td class="col2">1x</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">5, 6</td>
+                            <td class="col1">GFP + WT</td>
+                            <td class="col2">1x</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">7, 8</td>
+                            <td class="col1">GFP + strain 3</td>
+                            <td class="col2">3x</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">9, 10</td>
+                            <td class="col1">GFP + strain 7</td>
+                            <td class="col2">3x</td>
+                        </tr>
+                        <tr class="row6">
+                            <td class="col0">11, 12</td>
+                            <td class="col1">GFP + WT</td>
+                            <td class="col2">3x</td>
+                        </tr>
+                        <tr class="row7">
+                            <td class="col0">13, 14</td>
+                            <td class="col1">GFP + strain 3</td>
+                            <td class="col2">5x</td>
+                        </tr>
+                        <tr class="row8">
+                            <td class="col0">15, 16</td>
+                            <td class="col1">GFP + strain 7</td>
+                            <td class="col2">5x</td>
+                        </tr>
+                        <tr class="row9">
+                            <td class="col0">17, 18</td>
+                            <td class="col1">GFP + WT</td>
+                            <td class="col2">5x</td>
+                        </tr>
+                        <tr class="row10">
+                            <td class="col0">19, 20</td>
+                            <td class="col1">WT (negative control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                        <tr class="row11">
+                            <td class="col0">21, 22</td>
+                            <td class="col1">GFP (positive control)</td>
+                            <td class="col2"></td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT36 TABLE [37646-37996] -->
+            </div>
+            <h3><a name="section10072016" id="section10072016">10/07/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Fluorescence microscopy 32 (Nikon C2+ confocal microscope): Wash-test B: GFP + spores (in solution)</strong> </p>
+                <p> –&gt; spores and WT show similar levels of fluorescence, probably autofluorescence </p>
+                <p> <strong>Sample preparation 33: Blocking-test A: spores milk powder + GFP (in solution)</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> solution centrifuged (2 min, 13,400 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> spores washed twice with PBS(1x) (2x 100µl)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> spores resuspended and incubated in milk powder(5%) (100µl, incubation time: see table below)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with GFP (5µl, 1,35g/ml, 90 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> washed with PBS(1x) (3x 100µl)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> resuspended in PBS(1x) (20 µl)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation on GOPTS-slide (4µl, 30 min, 4°C)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed and blocked with milk powder(5%) (30 min, 10 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with PBS(1x) </div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> rinsed with ddH2O</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> dried with compressed air</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with Mowiol DABCO (overnight)</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit37">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                            <td class="col2"><strong>Incubation time</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1, 2</td>
+                            <td class="col1">strain 3 + milk powder + GFP</td>
+                            <td class="col2">30 min</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">3, 4</td>
+                            <td class="col1">strain 3 + milk powder + GFP</td>
+                            <td class="col2">60 min</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">5, 6</td>
+                            <td class="col1">strain 7 + milk powder + GFP</td>
+                            <td class="col2">30 min</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">7, 8</td>
+                            <td class="col1">strain 7 + milk powder + GFP</td>
+                            <td class="col2">60 min</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">9, 10</td>
+                            <td class="col1">WT + milk powder + GFP (negative control)</td>
+                            <td class="col2">30 min</td>
+                        </tr>
+                        <tr class="row6">
+                            <td class="col0">11, 12</td>
+                            <td class="col1">WT + milk powder + GFP(negative control)</td>
+                            <td class="col2">60 min</td>
+                        </tr>
+                        <tr class="row7">
+                            <td class="col0">13, 14</td>
+                            <td class="col1">WT + milk powder (negative control)</td>
+                            <td class="col2">30 min</td>
+                        </tr>
+                        <tr class="row8">
+                            <td class="col0">15, 16</td>
+                            <td class="col1">WT + milk powder (negative control)</td>
+                            <td class="col2">60 min</td>
+                        </tr>
+                        <tr class="row9">
+                            <td class="col0">17, 18</td>
+                            <td class="col1">strain 3 + milk powder (negative control)</td>
+                            <td class="col2">30 min</td>
+                        </tr>
+                        <tr class="row10">
+                            <td class="col0">19, 20</td>
+                            <td class="col1">strain 3 + milk powder (negative control)</td>
+                            <td class="col2">60 min</td>
+                        </tr>
+                        <tr class="row11">
+                            <td class="col0">21, 22</td>
+                            <td class="col1">strain 7 + milk powder (negative control)</td>
+                            <td class="col2">30 min</td>
+                        </tr>
+                        <tr class="row12">
+                            <td class="col0">23, 24</td>
+                            <td class="col1">strain 7 + milk powder (negative control)</td>
+                            <td class="col2">60 min</td>
+                        </tr>
+                        <tr class="row13">
+                            <td class="col0">25, 26</td>
+                            <td class="col1">GFP (positive control)</td>
+                            <td class="col2">30 min</td>
+                        </tr>
+                    </table>
+                </div>
+                <!-- EDIT37 TABLE [38837-39543] -->
+            </div>
+        </div>
+        <!-- para_20-->
+    </div>
+    <!-- color -->
+    <div class="color2">
+        <div class="para_20">
+            <h3><a name="section10082016" id="section10082016">10/08/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Fluorescence microscopy 33 (Nikon C2+ confocal microscope): Blocking-test A: spores + milk powder + GFP (in solution)</strong> </p>
+                <p> –&gt; spores and WT show similar levels of fluorescence </p>
+            </div>
+            <h3><a name="section10112016" id="section10112016">10/11/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Sample preparation 34: Blocking-test B: spores + BSA + GFP (in solution)</strong> </p>
+                <ol>
+                    <li class="level1">
+                        <div class="li"> solution centrifuged (2 min, 13,400 rpm)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> spores (5,000,000) washed with PBS(1x) (3x 100µl)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> spores resuspended and incubated in BSA(5%) (100µl, 60 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with GFP (50µl, 100µg/ml, 60 min)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> washed with TBST(0.05%) (5x 100µl)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> resuspended in PBS(1x) (50 µl)</div>
+                    </li>
+                    <li class="level1">
+                        <div class="li"> incubation with Mowiol DABCO (overnight)</div>
+                    </li>
+                </ol>
+                <div class="table sectionedit38">
+                    <table class="inline">
+                        <tr class="row0">
+                            <td class="col0"><strong>Spot No.</strong></td>
+                            <td class="col1"><strong>Sample</strong></td>
+                        </tr>
+                        <tr class="row1">
+                            <td class="col0">1, 2</td>
+                            <td class="col1">strain 3 + BSA + GFP</td>
+                        </tr>
+                        <tr class="row2">
+                            <td class="col0">3, 4</td>
+                            <td class="col1">strain 7 + BSA + GFP</td>
+                        </tr>
+                        <tr class="row3">
+                            <td class="col0">5, 6</td>
+                            <td class="col1">WT + BSA + GFP (negative control)</td>
+                        </tr>
+                        <tr class="row4">
+                            <td class="col0">7, 8</td>
+                            <td class="col1">WT + BSA (negative control)</td>
+                        </tr>
+                        <tr class="row5">
+                            <td class="col0">9, 10</td>
+                            <td class="col1">strain 3 + milk powder (negative control)</td>
+                        </tr>
+                        <tr class="row6">
+                            <td class="col0">11, 12</td>
+                            <td class="col1">strain 7 + milk powder (negative control)</td>
+                        </tr>
+                        <tr class="row7">
+                            <td class="col0">13, 14</td>
+                            <td class="col1">GFP (positive control)</td>
+                        </tr>
+                    </table>
+                </div>
+            </div>
+            <h3><a name="section10122016" id="section10122016">10/12/2016</a></h3>
+            <div class="level4">
+                <p> <strong>Fluorescence microscopy 34 (Nikon C2+ confocal microscope): Blocking-test B: spores + BSA + GFP (in solution)</strong> </p>
+                <p> –&gt; too little spores </p>
+                <div class="tags"><span>
+	<a href="/igem2016/doku.php?id=tag:lab_book" class="wikilink1" title="tag:lab_book" rel="tag">lab book</a>
+</span></div>
+            </div>
+        </div>
+        <!-- para_20-->
+    </div>
+    <!-- color -->
+</body>
 </html>
 {{Freiburg/Footer}}