Revision as of 16:15, 17 October 2016

Our Protocols

Group 4

Members: Kevin, Christian, Vivi, Nathalie

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/Styles}} {{Freiburg/Menubar}} {{Freiburg/Away}} {{Freiburg/Main}}{{Freiburg/Top}}
-{{Freiburg/Menubar}}
-{{Freiburg/Away}}
-{{Freiburg/Main}}
 <html>
@@ Line 18: / Line 15: @@
          h3 {
              font-size: 36px;
-            padding: 20px;
+             padding: 20px 0px;
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@@ Line 30: / Line 28: @@
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-        .para_20 {
-            padding: 5% 15%;
-        }
          .image img {
              margin: 20px;
@@ Line 52: / Line 47: @@
 <body>
-     <div class="img_menu">
+     <div class="color1">
-        <h5 style="text-align: center"> Lab Journals </h5>
-        <center>
+            <div class="img_menu">
-            <div class="image">
+                <h5 style="text-align: center">Our Protocols    </h5>
-                <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>
+                <center>
-                <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>
+                    <div class="image">
-                <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/NotebookCloning"> <img class="imga" src="https://static.igem.org/mediawiki/2016/1/14/T--Freiburg--CloningLab.png" alt="Smiley face"></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/NotebookSpores"> <img class="imgb" src="https://static.igem.org/mediawiki/2016/9/93/T--Freiburg--SporesLab.png" alt="Smiley face"></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>
+                        <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>
-            </div>
+                        <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>
-        </center>
+                        <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>
+                    </div>
+                </center>
+        </div>
      </div>
+<h1 class="sectionedit1"><a name="group_4" id="group_4">Group 4</a></h1>
-<h5 class="sectionedit1">Group 4</h5>
 <div class="level1">
 <p>
-<strong>Members:Kevin, Christian, Vivi, Nathalie</strong><br/>
+<strong>Members: Kevin, Christian, Vivi, Nathalie</strong><br/>
 </p>
@@ Line 103: / Line 103: @@
 	</tr>
 </table></div>
-<!-- EDIT2 TABLE [127-426] -->
+<!-- EDIT2 TABLE [128-427] -->
 </div>
@@ Line 114: / Line 114: @@
 <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.
+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>
@@ Line 156: / Line 156: @@
 <p>
-<br/>
 <strong>Sample preparation 1: GOPTS + anti-GFP nanobody + B52-B56</strong>
 </p>
@@ Line 171: / Line 169: @@
 <p>
-<br/>
 <strong>Sample preparation 1: GOPTS + anti-GFP nanobody + B52-B56</strong>
 </p>
@@ Line 213: / Line 209: @@
 	</tr>
 </table></div>
-<!-- EDIT3 TABLE [1613-1773] -->
+<!-- EDIT3 TABLE [1598-1758] -->
-<p>
-<br/>
-</p>
 </div>
@@ Line 225: / Line 216: @@
 <p>
-<br/>
 <strong>Sample preparation 1: GOPTS + anti-GFP nanobody + B52-B56</strong> <br/>
@@ Line 240: / Line 229: @@
 <p>
-–&gt; pictures inconclusive:pictures inconclusive: wrong coverslips (second glass slide), no sufficient controls <br/>
+–&gt; pictures inconclusive: wrong coverslips (second glass slide), no sufficient controls <br/>
 </p>
@@ Line 258: / Line 247: @@
 </li>
 </ol>
-<p>
-<br/>
-</p>
 <div class="table sectionedit4"><table class="inline">
 	<tr class="row0">
@@ Line 289: / Line 273: @@
 	</tr>
 </table></div>
-<!-- EDIT4 TABLE [2341-2570] -->
+<!-- EDIT4 TABLE [2287-2516] -->
-<p>
-<br/>
-</p>
 </div>
@@ Line 301: / Line 280: @@
 <p>
-<br/>
 <strong>Sample preparation</strong>
 </p>
@@ Line 320: / Line 297: @@
 <p>
-<br/>
 <strong>Sample preparation 2: GOPTS + anti-GFP nanobody + B52-56</strong>
 </p>
@@ Line 332: / Line 307: @@
 <p>
-<br/>
 –&gt; <strong><em class="u">FACS-analysis has shown that only B53 and B54 show fluorescence linked to GFP</em></strong>
 </p>
@@ Line 343: / Line 316: @@
 <p>
-<br/>
 <strong>Preparation of samples 3: B53-B55</strong>
 </p>
@@ Line 369: / Line 340: @@
 	</tr>
 </table></div>
-<!-- EDIT5 TABLE [3224-3273] -->
+<!-- EDIT5 TABLE [3152-3201] -->
-<p>
-<br/>
-</p>
 <p>
 <strong>Fluorescence microscopy 2 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + B52-56</strong>
@@ Line 405: / Line 371: @@
 <p>
-<br/>
 <strong>Sample preparation 4: GOPTS + anti-GFP nanobody + B53/B54</strong>
 </p>
@@ Line 420: / Line 384: @@
 <p>
-<br/>
 <strong>Sample preparation 4: GOPTS + anti-GFP nanobody + B53/B54</strong>
 </p>
@@ Line 460: / Line 422: @@
 	</tr>
 </table></div>
-<!-- EDIT6 TABLE [4077-4329] -->
+<!-- EDIT6 TABLE [3988-4240] -->
-<p>
-<br/>
-</p>
 </div>
@@ Line 472: / Line 429: @@
 <p>
-<br/>
 <strong>Fluorescence microscopy 4 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + B53/B54</strong>
 </p>
@@ Line 487: / Line 442: @@
 <p>
-<br/>
 <strong>Sample preparation 5:  GOPTS + anti-GFP nanobody + B53-56/GFP</strong>
 </p>
@@ Line 551: / Line 504: @@
 	</tr>
 </table></div>
-<!-- EDIT7 TABLE [4943-5475] -->
+<!-- EDIT7 TABLE [4840-5372] -->
-<p>
-<br/>
-</p>
 </div>
@@ Line 563: / Line 511: @@
 <p>
-<br/>
 <strong>Fluorescence microscopy 5 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + B53-56/GFP</strong>
 </p>
 <p>
-–&gt; only fluorescence detectable on spots 6 and 8
+–&gt; spores have not been bound
-</p>
-<p>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:8.8.16-_alexanb_b54.jpg" class="media" title="labprotocol:8.8.16-_alexanb_b54.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:8.8.16-_alexanb_b54.jpg" class="medialeft" align="left" alt="" width="200" /></a>1.)alexaNB+B54,
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:8.8.16-gopts_b54_fluo.jpg" class="media" title="labprotocol:8.8.16-gopts_b54_fluo.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:8.8.16-gopts_b54_fluo.jpg" class="medialeft" align="left" alt="" width="200" /></a>2.)GOPTS+B54,
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:8.8.16-gopts_gfp.jpg" class="media" title="labprotocol:8.8.16-gopts_gfp.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:8.8.16-gopts_gfp.jpg" class="medialeft" align="left" alt="" width="200" /></a>3)GOPTS+GFP,
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:8.8.16_-_alexananobody_b53_fluo.jpg" class="media" title="labprotocol:8.8.16_-_alexananobody_b53_fluo.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:8.8.16_-_alexananobody_b53_fluo.jpg" class="medialeft" align="left" alt="" width="200" /></a>4)alexaNB+B53
 </p>
@@ Line 585: / Line 524: @@
 <p>
-<br/>
 <strong>Sample preparation 6: anti-GFP nanobody + spores (WT, B53, B54, B56) (in solution)</strong>
 </p>
@@ Line 632: / Line 569: @@
 	</tr>
 </table></div>
-<!-- EDIT8 TABLE [6322-6594] -->
+<!-- EDIT8 TABLE [5936-6208] -->
-<p>
-<br/>
-</p>
 <p>
 <strong>Fluorescence microscopy 6 (Nikon C2+ confocal microscope): anti-GFP nanobody + spores (WT, B53, B54, B56) in solution</strong>
@@ Line 652: / Line 584: @@
 <p>
-<br/>
 <strong>Sample preparation 7: GOPTS + anti-GFP nanobody + B53/B54</strong>
 </p>
@@ Line 701: / Line 631: @@
 	</tr>
 </table></div>
-<!-- EDIT9 TABLE [7293-7490] -->
+<!-- EDIT9 TABLE [6899-7096] -->
-<p>
-<br/>
-</p>
 <p>
 <strong> Washing test A: GOPTS + anti-GFP nanobody + B53</strong>
@@ Line 721: / Line 646: @@
 <p>
-<br/>
 <strong>Washing test A: GOPTS + anti-GFP nanobody + B53</strong>
-<br/>
 </p>
 <ol>
@@ Line 739: / Line 660: @@
 </li>
 </ol>
-<p>
-<br/>
-</p>
 </div>
@@ Line 751: / Line 667: @@
 <p>
-<br/>
 <strong>Washing test A: GOPTS + anti-GFP nanobody + B53</strong>
 </p>
@@ Line 765: / Line 679: @@
 </li>
 </ol>
-<p>
-<br/>
-</p>
 <div class="table sectionedit10"><table class="inline">
 	<tr class="row0">
@@ Line 790: / Line 699: @@
 	</tr>
 </table></div>
-<!-- EDIT10 TABLE [8140-8546] -->
+<!-- EDIT10 TABLE [7723-8129] -->
-<p>
-<br/>
-</p>
 </div>
-<h4><a name="section17082016" id="section17082016">17/08/2016</a></h4>
+<h4><a name="section08172016" id="section08172016">08/17/2016</a></h4>
 <div class="level4">
 <p>
-<br/>
 <strong>Fluorescence microscopy 7 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + B53/B54</strong>
 </p>
 <p>
-–&gt; spores have not been bound to nanobody
+–&gt; spores have not bound to nanobody
-<br/>
 </p>
@@ Line 818: / Line 718: @@
 <p>
-–&gt;spores have not bound to BSA, picture of BSA + B53 must have been a mistake
+–&gt; spores have not bound to BSA
-<br/>
 </p>
@@ Line 829: / Line 727: @@
 <p>
-<br/>
 <strong>Sample preparation 8: GOPTS + anti-GFP nanobody + GFP</strong>
 </p>
@@ Line 876: / Line 772: @@
 	</tr>
 </table></div>
-<!-- EDIT11 TABLE [9421-9665] -->
+<!-- EDIT11 TABLE [8932-9176] -->
-<p>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:21.8.16-freigem_2016_1.1_21.08.1ulgfpauf_gpots.jpg" class="media" title="labprotocol:21.8.16-freigem_2016_1.1_21.08.1ulgfpauf_gpots.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:21.8.16-freigem_2016_1.1_21.08.1ulgfpauf_gpots.jpg" class="medialeft" align="left" alt="" width="200" /></a>1)1µlGFP auf GOPTS
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:21.8.16-freigem_2016_1.4_10s5ulgfpaufgopts.jpg" class="media" title="labprotocol:21.8.16-freigem_2016_1.4_10s5ulgfpaufgopts.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:21.8.16-freigem_2016_1.4_10s5ulgfpaufgopts.jpg" class="medialeft" align="left" alt="" width="200" /></a>2)5µlGFP auf GOPTS
-</p>
-<p>
-<br/>
-</p>
 </div>
@@ Line 893: / Line 779: @@
 <p>
-<br/>
 <strong>Fluorescence microscopy 8 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + GFP</strong>
 </p>
 <p>
-–&gt;
+–&gt; GFP not bound, GOPTS plates too old
-<br/>
 </p>
@@ Line 910: / Line 792: @@
 <p>
-<br/>
 <strong>Sample preparation 9: GOPTS + GFP</strong>
 </p>
@@ Line 933: / Line 813: @@
 	</tr>
 	<tr class="row1">
-		<td class="col0">1</td><td class="col1">GOPTS + GFP (1 µl, 10 min) </td>
+		<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>
+		<td class="col0">2</td><td class="col1">GOPTS + GFP (2 µl, 10 min)</td>
 	</tr>
 	<tr class="row3">
@@ Line 963: / Line 843: @@
 	</tr>
 </table></div>
-<!-- EDIT12 TABLE [10312-10657] -->
+<!-- EDIT12 TABLE [9664-10007] -->
-<p>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:24.8.16-1.1.2gopts_gfp_1ul-10min-10s_fl.jpg" class="media" title="labprotocol:24.8.16-1.1.2gopts_gfp_1ul-10min-10s_fl.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:24.8.16-1.1.2gopts_gfp_1ul-10min-10s_fl.jpg" class="medialeft" align="left" alt="" width="200" /></a>(1)
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:24.8.16-1.2gopts_gfp-1ul-10min_10s_fl.jpg" class="media" title="labprotocol:24.8.16-1.2gopts_gfp-1ul-10min_10s_fl.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:24.8.16-1.2gopts_gfp-1ul-10min_10s_fl.jpg" class="medialeft" align="left" alt="" width="200" /></a>(2)
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:24.8.16-1.3_gopts_5ulgfp_fl.jpg" class="media" title="labprotocol:24.8.16-1.3_gopts_5ulgfp_fl.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:24.8.16-1.3_gopts_5ulgfp_fl.jpg" class="medialeft" align="left" alt="" width="200" /></a>(3)
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:24.8.16-2.1_gopts_1ulgfp_20s_fl.jpg" class="media" title="labprotocol:24.8.16-2.1_gopts_1ulgfp_20s_fl.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:24.8.16-2.1_gopts_1ulgfp_20s_fl.jpg" class="medialeft" align="left" alt="" width="200" /></a>(4)
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:24.8.16-2.2_gopts_2ulgfp15min_20s_fl.jpg" class="media" title="labprotocol:24.8.16-2.2_gopts_2ulgfp15min_20s_fl.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:24.8.16-2.2_gopts_2ulgfp15min_20s_fl.jpg" class="medialeft" align="left" alt="" width="200" /></a>(5)
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:24.8.16-2.3_gopts_5ulgfp15min_20s_fl.jpg" class="media" title="labprotocol:24.8.16-2.3_gopts_5ulgfp15min_20s_fl.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:24.8.16-2.3_gopts_5ulgfp15min_20s_fl.jpg" class="medialeft" align="left" alt="" width="200" /></a>(6)
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:24.8.16-3.1_gopts_1ulgfp20min_20s_fl.jpg" class="media" title="labprotocol:24.8.16-3.1_gopts_1ulgfp20min_20s_fl.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:24.8.16-3.1_gopts_1ulgfp20min_20s_fl.jpg" class="medialeft" align="left" alt="" width="200" /></a>(7)
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:24.8.16-3.2_gopts_2ulgfp20min_20s_fl.jpg" class="media" title="labprotocol:24.8.16-3.2_gopts_2ulgfp20min_20s_fl.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:24.8.16-3.2_gopts_2ulgfp20min_20s_fl.jpg" class="medialeft" align="left" alt="" width="200" /></a>(8)
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:24.8.16-3.3_gopts_5ulgfp20min_20s_fl.jpg" class="media" title="labprotocol:24.8.16-3.3_gopts_5ulgfp20min_20s_fl.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:24.8.16-3.3_gopts_5ulgfp20min_20s_fl.jpg" class="medialeft" align="left" alt="" width="200" /></a>(9)
-</p>
-<p>
-<br/>
-</p>
 <p>
 <strong>Sample preparation 10: GOPTS + anti-GFP nanobody + B53</strong>
-<br/>
 </p>
 <ol>
@@ Line 1,008: / Line 869: @@
 </li>
 </ol>
-<p>
-<br/>
-</p>
 <div class="table sectionedit13"><table class="inline">
 	<tr class="row0">
@@ Line 1,033: / Line 889: @@
 	</tr>
 </table></div>
-<!-- EDIT13 TABLE [11730-11930] -->
+<!-- EDIT13 TABLE [10477-10677] -->
 <p>
-<br/>
 <strong>Washing test B: GOPTS + anti-GFP nanobody + B53</strong>
 </p>
@@ Line 1,061: / Line 915: @@
 </li>
 </ol>
-<p>
-<br/>
-</p>
 <div class="table sectionedit14"><table class="inline">
 	<tr class="row0">
@@ Line 1,101: / Line 950: @@
 	</tr>
 </table></div>
-<!-- EDIT14 TABLE [12400-13095] -->
+<!-- EDIT14 TABLE [11141-11836] -->
-<p>
-<br/>
-</p>
 </div>
-<h4><a name="section24082016" id="section24082016">24/08/2016</a></h4>
+<h4><a name="section08242016" id="section08242016">08/24/2016</a></h4>
 <div class="level4">
 <p>
-<br/>
 <strong>Fluorescence microscopy 9 (Nikon C2+ confocal microscope): GOPTS + GFP</strong>
 </p>
@@ Line 1,121: / Line 963: @@
 –&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>
+<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:24.8.16-1.1.2gopts_gfp_1ul-10min-10s_fl.jpg" class="media" title="labprotocol:24.8.16-1.1.2gopts_gfp_1ul-10min-10s_fl.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:24.8.16-1.1.2gopts_gfp_1ul-10min-10s_fl.jpg" class="medialeft" align="left" alt="" width="200" /></a>
+<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:24.8.16-1.2gopts_gfp-1ul-10min_10s_fl.jpg" class="media" title="labprotocol:24.8.16-1.2gopts_gfp-1ul-10min_10s_fl.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:24.8.16-1.2gopts_gfp-1ul-10min_10s_fl.jpg" class="medialeft" align="left" alt="" width="200" /></a>
+<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:24.8.16-1.3_gopts_5ulgfp_fl.jpg" class="media" title="labprotocol:24.8.16-1.3_gopts_5ulgfp_fl.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:24.8.16-1.3_gopts_5ulgfp_fl.jpg" class="medialeft" align="left" alt="" width="200" /></a>
+<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:24.8.16-2.1_gopts_1ulgfp_20s_fl.jpg" class="media" title="labprotocol:24.8.16-2.1_gopts_1ulgfp_20s_fl.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:24.8.16-2.1_gopts_1ulgfp_20s_fl.jpg" class="medialeft" align="left" alt="" width="200" /></a>
+<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:24.8.16-2.2_gopts_2ulgfp15min_20s_fl.jpg" class="media" title="labprotocol:24.8.16-2.2_gopts_2ulgfp15min_20s_fl.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:24.8.16-2.2_gopts_2ulgfp15min_20s_fl.jpg" class="medialeft" align="left" alt="" width="200" /></a>
+<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:24.8.16-2.3_gopts_5ulgfp15min_20s_fl.jpg" class="media" title="labprotocol:24.8.16-2.3_gopts_5ulgfp15min_20s_fl.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:24.8.16-2.3_gopts_5ulgfp15min_20s_fl.jpg" class="medialeft" align="left" alt="" width="200" /></a>
+<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:24.8.16-3.1_gopts_1ulgfp20min_20s_fl.jpg" class="media" title="labprotocol:24.8.16-3.1_gopts_1ulgfp20min_20s_fl.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:24.8.16-3.1_gopts_1ulgfp20min_20s_fl.jpg" class="medialeft" align="left" alt="" width="200" /></a>
+<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:24.8.16-3.2_gopts_2ulgfp20min_20s_fl.jpg" class="media" title="labprotocol:24.8.16-3.2_gopts_2ulgfp20min_20s_fl.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:24.8.16-3.2_gopts_2ulgfp20min_20s_fl.jpg" class="medialeft" align="left" alt="" width="200" /></a>
+<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:24.8.16-3.3_gopts_5ulgfp20min_20s_fl.jpg" class="media" title="labprotocol:24.8.16-3.3_gopts_5ulgfp20min_20s_fl.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:24.8.16-3.3_gopts_5ulgfp20min_20s_fl.jpg" class="medialeft" align="left" alt="" width="200" /></a>
+</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>
@@ Line 1,140: / Line 1,014: @@
 </div>
-<h4><a name="section25082016" id="section25082016">25/08/2016</a></h4>
+<h4><a name="section08252016" id="section08252016">08/25/2016</a></h4>
 <div class="level4">
 <p>
-<br/>
 <strong>Blocking test C: GOPTS + BSA/milk powder + GFP</strong>
 </p>
@@ Line 1,178: / Line 1,050: @@
 	</tr>
 </table></div>
-<!-- EDIT15 TABLE [14062-14183] -->
+<!-- EDIT15 TABLE [13676-13797] -->
-<p>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:25.8.16-blockingtest_gfp_control.jpg" class="media" title="labprotocol:25.8.16-blockingtest_gfp_control.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:25.8.16-blockingtest_gfp_control.jpg" class="medialeft" align="left" alt="" width="200" /></a>(1)
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:25.8.16-blockingtest_bsa_gfp_20s_4x.jpg" class="media" title="labprotocol:25.8.16-blockingtest_bsa_gfp_20s_4x.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:25.8.16-blockingtest_bsa_gfp_20s_4x.jpg" class="medialeft" align="left" alt="" width="200" /></a>(2)
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:25.8.16-blockingtest_milchpuler_gfp_20s_4x.jpg" class="media" title="labprotocol:25.8.16-blockingtest_milchpuler_gfp_20s_4x.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:25.8.16-blockingtest_milchpuler_gfp_20s_4x.jpg" class="medialeft" align="left" alt="" width="200" /></a>(3)
-</p>
 <p>
 <strong>Contamination Test D: BSA, PBS Mowiol DABCO</strong>
@@ Line 1,191: / Line 1,057: @@
 <p>
 –&gt; no contamination/pollution detected
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:26.8.16-contamination-test-pbs_dabco_6s_20x.jpg" class="media" title="labprotocol:26.8.16-contamination-test-pbs_dabco_6s_20x.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:26.8.16-contamination-test-pbs_dabco_6s_20x.jpg" class="mediacenter" alt="" width="200" /></a>PBS+DABCO
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:26.8.16-contamination-_test-bsa_dabco_10s_60x.jpg" class="media" title="labprotocol:26.8.16-contamination-_test-bsa_dabco_10s_60x.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:26.8.16-contamination-_test-bsa_dabco_10s_60x.jpg" class="mediacenter" alt="" width="200" /></a>
 </p>
@@ Line 1,232: / Line 1,096: @@
 	</tr>
 </table></div>
-<!-- EDIT16 TABLE [14951-15139] -->
+<!-- EDIT16 TABLE [14206-14394] -->
-<p>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:26.8.16--gopts_20s_4x.jpg" class="media" title="labprotocol:26.8.16--gopts_20s_4x.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:26.8.16--gopts_20s_4x.jpg" class="mediacenter" alt="" width="200" /></a>GOPTS(neg Control)
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:26.8.16-gopts_mcherry_900s_incubationtime_10s_4x.jpg" class="media" title="labprotocol:26.8.16-gopts_mcherry_900s_incubationtime_10s_4x.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:26.8.16-gopts_mcherry_900s_incubationtime_10s_4x.jpg" class="mediacenter" alt="" width="200" /></a>GOPTS+mCherry 15 min
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:26.8.16-mcherry_eingetrocknet.jpg" class="media" title="labprotocol:26.8.16-mcherry_eingetrocknet.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:26.8.16-mcherry_eingetrocknet.jpg" class="medialeft" align="left" alt="" width="200" /></a>mCherry eingetrocknet (pos control)
-</p>
-<p>
-<br/>
-</p>
 <p>
 <strong>Sample preparation 12: GOPTS + anti-GFP nanobody + GFP</strong>
@@ Line 1,289: / Line 1,142: @@
 	</tr>
 </table></div>
-<!-- EDIT17 TABLE [15880-16078] -->
+<!-- EDIT17 TABLE [14873-15071] -->
-<p>
-<br/>
-</p>
 </div>
@@ Line 1,301: / Line 1,149: @@
 <p>
-<br/>
 <strong>Fluorescence microscopy C (Nikon C2+ confocal microscope): Blocking test: GOPTS + BSA/milkpowder + GFP</strong>
 </p>
@@ Line 1,315: / Line 1,161: @@
 <p>
-–&gt;no clear mCherry-signal, too weak (quantum yield of mCherry about 1/3 of that of GFP)?
+–&gt; no mCherry-signal, too weak (quantum yield of mCherry about 1/3 of that of GFP)
 </p>
@@ Line 1,323: / Line 1,169: @@
 <p>
-–&gt;GFP has not bound to nanobody
+–&gt; GFP has not bound to nanobody
-</p>
-<p>
-<br/>
 </p>
 </div>
-<h4><a name="section30082016" id="section30082016">30/08/2016</a></h4>
+<h4><a name="section08302016" id="section08302016">08/30/2016</a></h4>
 <div class="level4">
 <p>
-<br/>
 <strong>Sample preparation 13: GOPTS + nanobody + GFP</strong>
 </p>
@@ Line 1,383: / Line 1,222: @@
 	</tr>
 </table></div>
-<!-- EDIT18 TABLE [17067-17271] -->
+<!-- EDIT18 TABLE [16042-16246] -->
-<p>
-<br/>
-</p>
 </div>
@@ Line 1,395: / Line 1,229: @@
 <p>
-<br/>
 <strong>Fluorescence microscopy 13 (Nikon C2+ confocal microscope): GOPTS + nanobody + GFP</strong>
 </p>
@@ Line 1,402: / Line 1,234: @@
 <p>
 –&gt; no GFP signal detectable
-<br/>
 </p>
@@ Line 1,412: / Line 1,242: @@
 <p>
-<br/>
 <strong>Sample preparation 14: GOPTS + GFP</strong>
 </p>
@@ Line 1,430: / Line 1,258: @@
 </li>
 </ol>
-<p>
-<br/>
-</p>
 <div class="table sectionedit19"><table class="inline">
 	<tr class="row0">
@@ Line 1,452: / Line 1,275: @@
 	</tr>
 </table></div>
-<!-- EDIT19 TABLE [17745-17903] -->
+<!-- EDIT19 TABLE [16703-16861] -->
 <p>
 <em class="u">Mowiol was produced due to this protocol:</em> <a href="https://www.carlroth.com/downloads/ba/en/0/BA_0713_EN.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/>
@@ Line 1,460: / Line 1,283: @@
 </div>
-<h4><a name="section090716" id="section090716">09/07/16</a></h4>
+<h4><a name="section09072016" id="section09072016">09/07/2016</a></h4>
 <div class="level4">
 <p>
-<br/>
 <strong>Fluorescence microscopy 14 (Nikon C2+ confocal microscope): GOPTS + GFP</strong>
 </p>
@@ Line 1,474: / Line 1,295: @@
 <p>
-<strong>Fluorescence microscopy 14 (Nikon C2+ confocal microscope):</strong>
+<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:spot_4_4x_20s.jpg" class="media" title="labprotocol:spot_4_4x_20s.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:spot_4_4x_20s.jpg" class="medialeft" align="left" alt="" width="200" /></a>
-</p>
+<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:9.9.16_spoting_maske_test_positivkontrole_3_-_100_.jpg" class="media" title="labprotocol:9.9.16_spoting_maske_test_positivkontrole_3_-_100_.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:9.9.16_spoting_maske_test_positivkontrole_3_-_100_.jpg" class="medialeft" align="left" alt="" width="200" /></a>
-<p>
 <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>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:9.9.16_spot_3_4x_20s_2_mit_skala_0_1mm.jpg" class="media" title="labprotocol:9.9.16_spot_3_4x_20s_2_mit_skala_0_1mm.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:9.9.16_spot_3_4x_20s_2_mit_skala_0_1mm.jpg" class="medialeft" align="left" alt="" width="200" /></a>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:9.9.16_spot_3_4x_20s_2_mit_skala_0_1mm.jpg" class="media" title="labprotocol:9.9.16_spot_3_4x_20s_2_mit_skala_0_1mm.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:9.9.16_spot_3_4x_20s_2_mit_skala_0_1mm.jpg" class="medialeft" align="left" alt="" width="200" /></a>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:9.9.16_spot_4_4x_20s_2_.jpg" class="media" title="labprotocol:9.9.16_spot_4_4x_20s_2_.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:9.9.16_spot_4_4x_20s_2_.jpg" class="medialeft" align="left" alt="" width="200" /></a>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:9.9.16_spot_5.2_4x_20s_2_.jpg" class="media" title="labprotocol:9.9.16_spot_5.2_4x_20s_2_.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:9.9.16_spot_5.2_4x_20s_2_.jpg" class="medialeft" align="left" alt="" width="200" /></a>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:9.9.16_spoting_maske_test_positivkontrole_3_-_100_.jpg" class="media" title="labprotocol:9.9.16_spoting_maske_test_positivkontrole_3_-_100_.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:9.9.16_spoting_maske_test_positivkontrole_3_-_100_.jpg" class="medialeft" align="left" alt="" width="200" /></a>
 <br/>
-</p>
-<p>
-Spot 1, 3, 4, 5, Positive control
 </p>
@@ Line 1,502: / Line 1,318: @@
 <p>
-<br/>
 <strong>Sample preparation 16: GOPTS + B53</strong>
 </p>
@@ Line 1,522: / Line 1,336: @@
 <div class="table sectionedit20"><table class="inline">
 	<tr class="row0">
-		<th class="col0">Spot No.</th><th class="col1">Sample</th>
+		<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>
+		<td class="col0">1,2,3</td><td class="col1">GOPTS + B53</td>
 	</tr>
 	<tr class="row2">
@@ Line 1,540: / Line 1,354: @@
 	</tr>
 </table></div>
-<!-- EDIT20 TABLE [18887-19028] -->
+<!-- EDIT20 TABLE [17589-17737] -->
 </div>
@@ Line 1,547: / Line 1,361: @@
 <p>
-<br/>
 <strong>Sample preparation 17: GOPTS + anti-GFP nanobody(Alexa647) + GFP</strong>
 </p>
@@ Line 1,593: / Line 1,405: @@
 	</tr>
 </table></div>
-<!-- EDIT21 TABLE [19539-19745] -->
+<!-- EDIT21 TABLE [18244-18450] -->
-<p>
-<br/>
-</p>
 <p>
 <strong>Fluorescence microscopy 16 (Nikon C2+ confocal microscope): GOPTS + B53</strong>
@@ Line 1,605: / Line 1,412: @@
 <p>
 –&gt; no spores detectable
-</p>
-<p>
-<br/>
-<br/>
 </p>
 </div>
-<h4><a name="section091616" id="section091616">09/16/16</a></h4>
+<h4><a name="section09162016" id="section09162016">09/16/2016</a></h4>
 <div class="level4">
 <p>
-<br/>
 <strong>Fluorescence microscopy 17 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + GFP</strong>
-–&gt; GFP signal: GFP has bound to nanobody
 </p>
 <p>
-<br/>
+–&gt; GFP signal: GFP has been bound to nanobody
-Spot 1-4: NB+GFP
-<br/>
-<br/>
-<a href="/igem2016/lib/exe/fetch.php?media=labprotocol:nb_auf_gfp_spot_1_4s_gain_3_4_bearbeitet_.jpg" class="media" title="labprotocol:nb_auf_gfp_spot_1_4s_gain_3_4_bearbeitet_.jpg"><img src="/igem2016/lib/exe/fetch.php?w=300&amp;media=labprotocol:nb_auf_gfp_spot_1_4s_gain_3_4_bearbeitet_.jpg" class="media" alt="" width="300" /></a><br/>
 </p>
 <p>
-<a href="/igem2016/lib/exe/fetch.php?media=labprotocol:nb_mit_gfp_20_s_spot_3_4x_bearbeitet_.jpg" class="media" title="labprotocol:nb_mit_gfp_20_s_spot_3_4x_bearbeitet_.jpg"><img src="/igem2016/lib/exe/fetch.php?w=300&amp;media=labprotocol:nb_mit_gfp_20_s_spot_3_4x_bearbeitet_.jpg" class="media" alt="" width="300" /></a><br/>
+<a href="/igem2016/lib/exe/fetch.php?media=labprotocol:nb_auf_gfp_spot_1_4s_gain_3_4_bearbeitet_.jpg" class="media" title="labprotocol:nb_auf_gfp_spot_1_4s_gain_3_4_bearbeitet_.jpg"><img src="/igem2016/lib/exe/fetch.php?w=300&amp;media=labprotocol:nb_auf_gfp_spot_1_4s_gain_3_4_bearbeitet_.jpg" class="media" alt="" width="300" /></a>
+<a href="/igem2016/lib/exe/fetch.php?media=labprotocol:nb_mit_gfp_20_s_spot_3_4x_bearbeitet_.jpg" class="media" title="labprotocol:nb_mit_gfp_20_s_spot_3_4x_bearbeitet_.jpg"><img src="/igem2016/lib/exe/fetch.php?w=300&amp;media=labprotocol:nb_mit_gfp_20_s_spot_3_4x_bearbeitet_.jpg" class="media" alt="" width="300" /></a>
-</p>
+<a href="/igem2016/lib/exe/fetch.php?media=labprotocol:nb_mit_gfp_4s_gain_2_8_spot_2_4x_bearbeitet_.jpg" class="media" title="labprotocol:nb_mit_gfp_4s_gain_2_8_spot_2_4x_bearbeitet_.jpg"><img src="/igem2016/lib/exe/fetch.php?w=300&amp;media=labprotocol:nb_mit_gfp_4s_gain_2_8_spot_2_4x_bearbeitet_.jpg" class="media" alt="" width="300" /></a>
+<a href="/igem2016/lib/exe/fetch.php?media=labprotocol:nb_mit_gfp_spot_4_20_s_4x_bearbeitet_.jpg" class="media" title="labprotocol:nb_mit_gfp_spot_4_20_s_4x_bearbeitet_.jpg"><img src="/igem2016/lib/exe/fetch.php?w=300&amp;media=labprotocol:nb_mit_gfp_spot_4_20_s_4x_bearbeitet_.jpg" class="media" alt="" width="300" /></a>
-<p>
+<a href="/igem2016/lib/exe/fetch.php?media=labprotocol:gfp_auf_milk_4s_gain_3_4_bearbeitet_.jpg" class="media" title="labprotocol:gfp_auf_milk_4s_gain_3_4_bearbeitet_.jpg"><img src="/igem2016/lib/exe/fetch.php?w=300&amp;media=labprotocol:gfp_auf_milk_4s_gain_3_4_bearbeitet_.jpg" class="media" alt="" width="300" /></a>
-<a href="/igem2016/lib/exe/fetch.php?media=labprotocol:nb_mit_gfp_4s_gain_2_8_spot_2_4x_bearbeitet_.jpg" class="media" title="labprotocol:nb_mit_gfp_4s_gain_2_8_spot_2_4x_bearbeitet_.jpg"><img src="/igem2016/lib/exe/fetch.php?w=300&amp;media=labprotocol:nb_mit_gfp_4s_gain_2_8_spot_2_4x_bearbeitet_.jpg" class="media" alt="" width="300" /></a><br/>
+<a href="/igem2016/lib/exe/fetch.php?media=labprotocol:nb_auf_milk_20s_4x_bearbeitet_.jpg" class="media" title="labprotocol:nb_auf_milk_20s_4x_bearbeitet_.jpg"><img src="/igem2016/lib/exe/fetch.php?w=300&amp;media=labprotocol:nb_auf_milk_20s_4x_bearbeitet_.jpg" class="media" alt="" width="300" /></a>
+<a href="/igem2016/lib/exe/fetch.php?media=labprotocol:gfp_auf_gopts_20_s_4x_bearbeitet_.jpg" class="media" title="labprotocol:gfp_auf_gopts_20_s_4x_bearbeitet_.jpg"><img src="/igem2016/lib/exe/fetch.php?w=300&amp;media=labprotocol:gfp_auf_gopts_20_s_4x_bearbeitet_.jpg" class="media" alt="" width="300" /></a>
-</p>
+<a href="/igem2016/lib/exe/fetch.php?media=labprotocol:nur_gopts_20s_4x_bearbeitet_.jpg" class="media" title="labprotocol:nur_gopts_20s_4x_bearbeitet_.jpg"><img src="/igem2016/lib/exe/fetch.php?w=300&amp;media=labprotocol:nur_gopts_20s_4x_bearbeitet_.jpg" class="media" alt="" width="300" /></a>
-<p>
-<a href="/igem2016/lib/exe/fetch.php?media=labprotocol:nb_mit_gfp_spot_4_20_s_4x_bearbeitet_.jpg" class="media" title="labprotocol:nb_mit_gfp_spot_4_20_s_4x_bearbeitet_.jpg"><img src="/igem2016/lib/exe/fetch.php?w=300&amp;media=labprotocol:nb_mit_gfp_spot_4_20_s_4x_bearbeitet_.jpg" class="media" alt="" width="300" /></a><br/>
 <br/>
-<br/>
-<strong>Spot 5 GFP on milkpowder</strong>
-→ negative Control  - GFP Signal only in glass scratch
-<br/>
-<br/>
-<a href="/igem2016/lib/exe/fetch.php?media=labprotocol:gfp_auf_milk_4s_gain_3_4_bearbeitet_.jpg" class="media" title="labprotocol:gfp_auf_milk_4s_gain_3_4_bearbeitet_.jpg"><img src="/igem2016/lib/exe/fetch.php?w=300&amp;media=labprotocol:gfp_auf_milk_4s_gain_3_4_bearbeitet_.jpg" class="media" alt="" width="300" /></a><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>
@@ Line 1,669: / Line 1,456: @@
 <br/>
-Spot 6 NB on MilkPowder → negative Control<br/>
-<a href="/igem2016/lib/exe/fetch.php?media=labprotocol:nb_auf_milk_20s_4x_bearbeitet_.jpg" class="media" title="labprotocol:nb_auf_milk_20s_4x_bearbeitet_.jpg"><img src="/igem2016/lib/exe/fetch.php?w=300&amp;media=labprotocol:nb_auf_milk_20s_4x_bearbeitet_.jpg" class="media" alt="" width="300" /></a><br/>
-</p>
-<p>
-<br/>
-<br/>
-Spot 7 GFP on GOPTS → positive control<br/>
-<a href="/igem2016/lib/exe/fetch.php?media=labprotocol:gfp_auf_gopts_20_s_4x_bearbeitet_.jpg" class="media" title="labprotocol:gfp_auf_gopts_20_s_4x_bearbeitet_.jpg"><img src="/igem2016/lib/exe/fetch.php?w=300&amp;media=labprotocol:gfp_auf_gopts_20_s_4x_bearbeitet_.jpg" class="media" alt="" width="300" /></a><br/>
-</p>
-<p>
-<br/>
-<br/>
-</p>
-<p>
-Spot 8 only GOPTS → negativ control<br/>
-<a href="/igem2016/lib/exe/fetch.php?media=labprotocol:nur_gopts_20s_4x_bearbeitet_.jpg" class="media" title="labprotocol:nur_gopts_20s_4x_bearbeitet_.jpg"><img src="/igem2016/lib/exe/fetch.php?w=300&amp;media=labprotocol:nur_gopts_20s_4x_bearbeitet_.jpg" class="media" alt="" width="300" /></a><br/>
 <br/>
@@ Line 1,707: / Line 1,465: @@
 </div>
-<h4><a name="section091716" id="section091716">09/17/16</a></h4>
+<h4><a name="section09172016" id="section09172016">09/17/2016</a></h4>
 <div class="level4">
 <p>
-<br/>
 <strong>Sample preparation 18: GOPTS + anti-GFP nanobody + GFP (spotting-mask)</strong>
 </p>
@@ Line 1,737: / Line 1,493: @@
 </li>
 </ol>
-<p>
-<br/>
-</p>
 </div>
-<h4><a name="section091816" id="section091816">09/18/16</a></h4>
+<h4><a name="section09182016" id="section09182016">09/18/2016</a></h4>
 <div class="level4">
 <p>
-<br/>
 <strong>Fluorescence microscopy 18 (Nikon C2+ confocal microscope): GOPTS + antiGFP-nanobody + GFP (spotting-mask)</strong>
 </p>
 <p>
-–&gt;no GFP signal detectable
+–&gt; no GFP signal detectable
 </p>
@@ Line 1,785: / Line 1,534: @@
 </li>
 </ol>
-<p>
-<br/>
-</p>
 </div>
-<h4><a name="section091916" id="section091916">09/19/16</a></h4>
+<h4><a name="section09192016" id="section09192016">09/19/2016</a></h4>
 <div class="level4">
 <p>
-<br/>
 <strong>Fluorescence microscopy 19 (Nikon C2+ confocal microscope): GOPTS + anti-GFP nanobody + GFP (spotting-mask)</strong>
 </p>
@@ Line 1,822: / Line 1,564: @@
 <p>
 –&gt; problem: no washing away of unbound nanobody
-</p>
-<p>
-<br/>
 </p>
 <div class="table sectionedit22"><table class="inline">
@@ Line 1,872: / Line 1,609: @@
 	</tr>
 </table></div>
-<!-- EDIT22 TABLE [22586-23371] -->
+<!-- EDIT22 TABLE [21203-21988] -->
-<p>
-<br/>
-</p>
 </div>
-<h4><a name="section092016" id="section092016">09/20/16</a></h4>
+<h4><a name="section09202016" id="section09202016">09/20/2016</a></h4>
 <div class="level4">
 <p>
-<br/>
 <strong>Sample preparation 21: anti-GFP nanobody + B53 (in solution)</strong>
 </p>
@@ Line 1,906: / Line 1,636: @@
 </li>
 </ol>
-<p>
-<br/>
-</p>
 <div class="table sectionedit23"><table class="inline">
 	<tr class="row0">
@@ Line 1,928: / Line 1,653: @@
 	</tr>
 </table></div>
-<!-- EDIT23 TABLE [23833-24082] -->
+<!-- EDIT23 TABLE [22443-22692] -->
-<p>
-<br/>
-</p>
 </div>
-<h4><a name="section092916" id="section092916">09/29/16</a></h4>
+<h4><a name="section09292016" id="section09292016">09/29/2016</a></h4>
 <div class="level4">
 <p>
-<br/>
 <strong>Sample preparation 22: GOPTS + GFP</strong>
 </p>
@@ Line 1,969: / Line 1,687: @@
 	</tr>
 </table></div>
-<!-- EDIT24 TABLE [24412-24467] -->
+<!-- EDIT24 TABLE [23013-23068] -->
 </div>
-<h4><a name="section093016" id="section093016">09/30/16</a></h4>
+<h4><a name="section09302016" id="section09302016">09/30/2016</a></h4>
 <div class="level4">
 <p>
-<br/>
 <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>No.</strong></td><td class="col1"><strong>Strain</strong></td>
+		<td class="col0"><strong>Strain No.</strong></td><td class="col1"><strong>Strain</strong></td>
 	</tr>
 	<tr class="row1">
-		<td class="col0">1</td><td class="col1">151:B. Subtilis CotG(3)</td>
+		<td class="col0">3</td><td class="col1">150: aGFPnano_HA_aHelix_cgeA</td>
 	</tr>
 	<tr class="row2">
-		<td class="col0">2</td><td class="col1">150 (6)</td>
+		<td class="col0">6</td><td class="col1">151: aGFPnano_HA_aHelix_cotG</td>
 	</tr>
 	<tr class="row3">
-		<td class="col0">3</td><td class="col1">159 (7)</td>
+		<td class="col0">7</td><td class="col1">159: aGFPnano_HA_G4S_CotZ</td>
 	</tr>
 	<tr class="row4">
-		<td class="col0">4</td><td class="col1">157 (9)</td>
+		<td class="col0">9</td><td class="col1">157: aGFPnano_HA_aHelix_CotZ</td>
 	</tr>
 </table></div>
-<!-- EDIT25 TABLE [24541-24625] -->
+<!-- EDIT25 TABLE [23141-23297] -->
 <p>
 <strong>Fluorescence microscopy 22 (Nikon C2+ confocal microscope): GOPTS + GFP</strong>
@@ Line 2,097: / Line 1,813: @@
 	</tr>
 </table></div>
-<!-- EDIT26 TABLE [25201-25999] -->
+<!-- EDIT26 TABLE [23872-24670] -->
 </div>
-<h4><a name="section100116" id="section100116">10/01/16</a></h4>
+<h4><a name="section10012016" id="section10012016">10/01/2016</a></h4>
 <div class="level4">
 <p>
-<br/>
+<strong>Sample preparation 23: GOPTS + GFP + spores</strong>
-<strong>Sample preparation 23: GOPTS + GFP + XXX</strong>
 </p>
@@ Line 2,114: / Line 1,828: @@
 <p>
-<strong>Sample preparation 24: GOPTS + GFP + XXX (in EtOH resuspended)</strong>
+<strong>Sample preparation 24: GOPTS + GFP + spores (in EtOH resuspended)</strong>
 </p>
 <ol>
@@ Line 2,160: / Line 1,874: @@
 	</tr>
 </table></div>
-<!-- EDIT27 TABLE [26599-26738] -->
+<!-- EDIT27 TABLE [25275-25414] -->
 </div>
-<h4><a name="section100216" id="section100216">10/02/16</a></h4>
+<h4><a name="section10022016" id="section10022016">10/02/2016</a></h4>
 <div class="level4">
 <p>
-<br/>
+<strong>Fluorescence microscopy 24 (Nikon C2+ confocal microscope): GOPTS + GFP + spores (strains 3,6,7,9,wt) (in EtOH resuspended)</strong>
-<strong>Fluorescence microscopy 24 (Nikon C2+ confocal microscope): GOPTS + GFP + XXX (in EtOH resuspended)</strong>
 </p>
 <p>
 –&gt; EtOH denaturated GFP: very weak signal. No spores detectable
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:slide_31_cgea-6_und_gfp_100x_2s.jpg" class="media" title="labprotocol:slide_31_cgea-6_und_gfp_100x_2s.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:slide_31_cgea-6_und_gfp_100x_2s.jpg" class="medialeft" align="left" alt="" width="200" /></a>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:slide_31_cgea-6_und_gfp_100x_2s_zweiter.jpg" class="media" title="labprotocol:slide_31_cgea-6_und_gfp_100x_2s_zweiter.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:slide_31_cgea-6_und_gfp_100x_2s_zweiter.jpg" class="medialeft" align="left" alt="" width="200" /></a>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:slide_31_cgea-6_und_gfp_100x_2s_zweiter_gfp_channel.jpg" class="media" title="labprotocol:slide_31_cgea-6_und_gfp_100x_2s_zweiter_gfp_channel.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:slide_31_cgea-6_und_gfp_100x_2s_zweiter_gfp_channel.jpg" class="medialeft" align="left" alt="" width="200" /></a>
 </p>
 <p>
-<strong>Sample preparation 25: GFP + XXX (in solution)</strong>
+<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) + XXX (4µl, 400,000/µl) (incubation time: see table below, 4°C)</div>
+<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>
@@ Line 2,203: / Line 1,912: @@
 <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 class="level1"><div class="li"> incubrnight)ation with Mowiol DABCO </div>
 </li>
 </ol>
@@ Line 2,247: / Line 1,956: @@
 	</tr>
 </table></div>
-<!-- EDIT28 TABLE [27670-28052] -->
+<!-- EDIT28 TABLE [26200-26582] -->
 </div>
-<h4><a name="section100316" id="section100316">10/03/16</a></h4>
+<h4><a name="section10032016" id="section10032016">10/03/2016</a></h4>
 <div class="level4">
 <p>
-<br/>
+<strong>Sample preparation 26: Incubation test: GFP + spores (in solution)</strong>
-<strong>Sample preparation 26: Incubation test: GFP + XXX (in solution)</strong>
 </p>
 <ol>
-<li class="level1"><div class="li"> incubation of GFP (4µl, 1,35g/ml) + XXX (4µl, 400,000/µl) (incubation time: see tables below, 4°C)</div>
+<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>
@@ Line 2,339: / Line 2,046: @@
 	</tr>
 </table></div>
-<!-- EDIT29 TABLE [28637-29334] -->
+<!-- EDIT29 TABLE [27172-27869] -->
 <p>
 Slide 2:
@@ Line 2,396: / Line 2,103: @@
 	</tr>
 </table></div>
-<!-- EDIT30 TABLE [29346-30043] -->
+<!-- EDIT30 TABLE [27881-28578] -->
 <p>
 Slide 3:
@@ Line 2,453: / Line 2,160: @@
 	</tr>
 </table></div>
-<!-- EDIT31 TABLE [30055-30760] -->
+<!-- EDIT31 TABLE [28590-29295] -->
 <p>
 <strong>Sample preparation 28: GFP-volume test B: GFP + strain 7 (in solution, washed)</strong>
@@ Line 2,525: / Line 2,232: @@
 	</tr>
 </table></div>
-<!-- EDIT32 TABLE [31327-31843] -->
+<!-- EDIT32 TABLE [29862-30378] -->
 <p>
 <strong>Sample preparation 29: Spore-amount test: GFP + strain 7 (in solution)</strong>
@@ Line 2,597: / Line 2,304: @@
 	</tr>
 </table></div>
-<!-- EDIT33 TABLE [32404-32991] -->
+<!-- EDIT33 TABLE [30939-31526] -->
 </div>
-<h4><a name="section100416" id="section100416">10/04/16</a></h4>
+<h4><a name="section10042016" id="section10042016">10/04/2016</a></h4>
 <div class="level4">
 <p>
-<br/>
+<strong>Fluorescence microscopy 24 (Nikon C2+ confocal microscope): GOPTS + GFP + spores (in EtOH resuspended)</strong>
+</p>
-<strong>Fluorescence microscopy 24 (Nikon C2+ confocal microscope): GOPTS + GFP + XXX (in EtOH resuspended)</strong>
+<p>
+–&gt; spore concentration too high, too many layers of spores
 </p>
 <p>
-–&gt;
+<strong>Fluorescence microscopy 25 (Nikon C2+ confocal microscope): GFP + spores (in solution)</strong>
 </p>
 <p>
-<strong>Fluorescence microscopy 25 (Nikon C2+ confocal microscope): GFP + XXX (in solution)</strong>
+–&gt; spore concentration too high, too many layers of spores
 </p>
 <p>
-–&gt;
+<strong>Fluorescence microscopy 26 (Nikon C2+ confocal microscope): Incubation test: GFP + spores (in solution)</strong>
 </p>
 <p>
-<strong>Fluorescence microscopy 26 (Nikon C2+ confocal microscope): Incubation test: GFP + XXX (in solution)</strong>
+–&gt; spore concentration too high, too many layers of spores
 </p>
 <p>
-–&gt;
+Slide 1:
 </p>
 <p>
+<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:161004-_slide37-stam7_mitgfpin_losung-gewaschen30min-1ms60xweis-a2.jpg" class="media" title="labprotocol:161004-_slide37-stam7_mitgfpin_losung-gewaschen30min-1ms60xweis-a2.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:161004-_slide37-stam7_mitgfpin_losung-gewaschen30min-1ms60xweis-a2.jpg" class="medialeft" align="left" alt="" width="200" /></a>
+<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:161004-_slide37-stam7_mitgfpin_losung-gewaschen30min-4s60x-a.jpg" class="media" title="labprotocol:161004-_slide37-stam7_mitgfpin_losung-gewaschen30min-4s60x-a.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:161004-_slide37-stam7_mitgfpin_losung-gewaschen30min-4s60x-a.jpg" class="medialeft" align="left" alt="" width="200" /></a>
+<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:161004-_slide37-stamm_9_mitgfpin_losung-gewaschen30min-1ms60x-a.jpg" class="media" title="labprotocol:161004-_slide37-stamm_9_mitgfpin_losung-gewaschen30min-1ms60x-a.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:161004-_slide37-stamm_9_mitgfpin_losung-gewaschen30min-1ms60x-a.jpg" class="medialeft" align="left" alt="" width="200" /></a>
+<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:161004-_slide37-stamm_9_mitgfpin_losung-gewaschen30min-2s60x-a.jpg" class="media" title="labprotocol:161004-_slide37-stamm_9_mitgfpin_losung-gewaschen30min-2s60x-a.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:161004-_slide37-stamm_9_mitgfpin_losung-gewaschen30min-2s60x-a.jpg" class="medialeft" align="left" alt="" width="200" /></a>
+<br/>
+</p>
+<p>
+.1) strain7 + GFP (30 min) (white field)
+.2) strain7 + GFP (30 min) (GFP-channel)
+.1) strain9 + GFP (30 min) (white field)
+.2) strain9 + GFP (30 min) (GFP-channel)
+<br/>
+<br/>
+<br/>
+<br/>
+<br/>
+<br/>
+<br/>
+<br/>
+Slide 2:
+<br/>
+<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:strain_7_1_5h_mit_gfp_spot_7_wl_100x_sec_pic.jpg" class="media" title="labprotocol:strain_7_1_5h_mit_gfp_spot_7_wl_100x_sec_pic.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:strain_7_1_5h_mit_gfp_spot_7_wl_100x_sec_pic.jpg" class="medialeft" align="left" alt="" width="200" /></a>
+<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:strain_7_1_5h_mit_gfp_spot_6_gfp_60x.jpg" class="media" title="labprotocol:strain_7_1_5h_mit_gfp_spot_6_gfp_60x.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:strain_7_1_5h_mit_gfp_spot_6_gfp_60x.jpg" class="medialeft" align="left" alt="" width="200" /></a>
+<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:strain_wt_1_5h_mit_gfp_spot_8_gfp100x.jpg" class="media" title="labprotocol:strain_wt_1_5h_mit_gfp_spot_8_gfp100x.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:strain_wt_1_5h_mit_gfp_spot_8_gfp100x.jpg" class="medialeft" align="left" alt="" width="200" /></a>
+<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:strain_wt_1_5h_mit_gfp_spot_8_wl_100x.jpg" class="media" title="labprotocol:strain_wt_1_5h_mit_gfp_spot_8_wl_100x.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:strain_wt_1_5h_mit_gfp_spot_8_wl_100x.jpg" class="medialeft" align="left" alt="" width="200" /></a>
+<br/>
+</p>
+<p>
+.1) strain 7 + GFP (90 min)(white field)
+.2) strain 7 + GFP (90 min)(GFP-channel)
+.1) strain WT + GFP (90 min)(white field)
+.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;
+–&gt; background fluorescence was too high
 </p>
@@ Line 2,642: / Line 2,412: @@
 <p>
-–&gt;
+–&gt; background fluorescence was too high
 </p>
@@ Line 2,650: / Line 2,420: @@
 <p>
-–&gt;
+–&gt; spores and WT show similar levels of fluorescence, probably autofluorescence
 </p>
 <p>
-<strong>Sample preparation 30: Wash-test A: GFP + XXX (in solution)</strong>
+<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:1005.st7.gfp.slide28.waschtest4x.100ms60xd.jpg" class="media" title="labprotocol:1005.st7.gfp.slide28.waschtest4x.100ms60xd.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:1005.st7.gfp.slide28.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.st7.gfp.slide28.waschtest4x.4s60xd2.jpg" class="media" title="labprotocol:1005.st7.gfp.slide28.waschtest4x.4s60xd2.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:1005.st7.gfp.slide28.waschtest4x.4s60xd2.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.slide28.waschtest4x.100ms60xc.jpg" class="media" title="labprotocol:1005.wt.gfp.slide28.waschtest4x.100ms60xc.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:1005.wt.gfp.slide28.waschtest4x.100ms60xc.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.slide28.waschtest4x.4s60xc.jpg" class="media" title="labprotocol:1005.wt.gfp.slide28.waschtest4x.4s60xc.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:1005.wt.gfp.slide28.waschtest4x.4s60xc.jpg" class="medialeft" align="left" alt="" width="200" /></a>
+<br/>
+</p>
+<p>
+) strain 7 + GFP (white field)
+) strain 7 + GFP (GFP-channel)
+) WT + GFP (white field)
+) 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) + XXX (25,000.000 spores)  (90 min, 4°C)</div>
+<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>
@@ Line 2,733: / Line 2,532: @@
 	</tr>
 </table></div>
-<!-- EDIT34 TABLE [34179-34720] -->
+<!-- EDIT34 TABLE [34545-35086] -->
 <p>
-<strong>Sample preparation 31: Wash-test B: GFP + XXX (in solution)</strong>
+<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) + XXX (25,000.000 spores)  (90 min, 4°C)</div>
+<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>
@@ Line 2,814: / Line 2,613: @@
 	</tr>
 </table></div>
-<!-- EDIT35 TABLE [35245-35786] -->
+<!-- EDIT35 TABLE [35617-36158] -->
 </div>
-<h4><a name="section100516" id="section100516">10/05/16</a></h4>
+<h4><a name="section10052016" id="section10052016">10/05/2016</a></h4>
 <div class="level4">
 <p>
-<br/>
+<strong>Fluorescence microscopy 30 (Nikon C2+ confocal microscope): Wash-test A: GFP + spores (strains 3,6,7,9) (in solution)</strong>
+</p>
-<strong>Fluorescence microscopy 30 (Nikon C2+ confocal microscope): Wash-test A: GFP + XXX (in solution)</strong>
+<p>
+–&gt; spore concentration too high, background fluorescence too high
 </p>
 <p>
-–&gt;
+<strong>Fluorescence microscopy 31 (Nikon C2+ confocal microscope): Wash-test B: GFP + spores (strains 3,6,7,9) (in solution)</strong>
 </p>
 <p>
-<strong>Fluorescence microscopy 31 (Nikon C2+ confocal microscope): Wash-test B: GFP + XXX (in solution)</strong>
+–&gt; indicates that strain 3 binds GFP
 </p>
 <p>
-–&gt;
+<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>
-<h4><a name="section100616" id="section100616">10/06/16</a></h4>
+<h4><a name="section10062016" id="section10062016">10/06/2016</a></h4>
 <div class="level4">
 <p>
-<br/>
 <strong>Sample preparation 32: Wash-test C: GFP + spores (in solution)</strong>
 </p>
@@ Line 2,910: / Line 2,732: @@
 	</tr>
 </table></div>
-<!-- EDIT36 TABLE [36574-36924] -->
+<!-- EDIT36 TABLE [37646-37996] -->
-<p>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:1006.str3.slide36.inc.test.4s60xj.jpg" class="media" title="labprotocol:1006.str3.slide36.inc.test.4s60xj.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:1006.str3.slide36.inc.test.4s60xj.jpg" class="medialeft" align="left" alt="" width="200" /></a>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:1006.str3_gfp.slide36.inc.test.1s60xf.jpg" class="media" title="labprotocol:1006.str3_gfp.slide36.inc.test.1s60xf.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:1006.str3_gfp.slide36.inc.test.1s60xf.jpg" class="medialeft" align="left" alt="" width="200" /></a>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:1006.str6.slide36.inc.test.100ms60xi.jpg" class="media" title="labprotocol:1006.str6.slide36.inc.test.100ms60xi.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:1006.str6.slide36.inc.test.100ms60xi.jpg" class="medialeft" align="left" alt="" width="200" /></a>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:1006.str6_gfp.slide36.inc.test.1s60xe.jpg" class="media" title="labprotocol:1006.str6_gfp.slide36.inc.test.1s60xe.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:1006.str6_gfp.slide36.inc.test.1s60xe.jpg" class="medialeft" align="left" alt="" width="200" /></a>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:1006.str9.slide36.inc.test.4s60xg.jpg" class="media" title="labprotocol:1006.str9.slide36.inc.test.4s60xg.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:1006.str9.slide36.inc.test.4s60xg.jpg" class="medialeft" align="left" alt="" width="200" /></a>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:1006.str9.slide36.inc.test.100ms60xg.jpg" class="media" title="labprotocol:1006.str9.slide36.inc.test.100ms60xg.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:1006.str9.slide36.inc.test.100ms60xg.jpg" class="medialeft" align="left" alt="" width="200" /></a>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:1006.wt.slide36.inc.test.100ms60xa.jpg" class="media" title="labprotocol:1006.wt.slide36.inc.test.100ms60xa.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:1006.wt.slide36.inc.test.100ms60xa.jpg" class="medialeft" align="left" alt="" width="200" /></a>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:1006.wt.slide36.inc.test.4s60xa.jpg" class="media" title="labprotocol:1006.wt.slide36.inc.test.4s60xa.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:1006.wt.slide36.inc.test.4s60xa.jpg" class="medialeft" align="left" alt="" width="200" /></a>
-</p>
 </div>
-<h4><a name="section100716" id="section100716">10/07/16</a></h4>
+<h4><a name="section10072016" id="section10072016">10/07/2016</a></h4>
 <div class="level4">
 <p>
-<br/>
 <strong>Fluorescence microscopy 32 (Nikon C2+ confocal microscope): Wash-test B: GFP + spores (in solution)</strong>
 </p>
 <p>
-–&gt;
+–&gt; spores and WT show similar levels of fluorescence, probably autofluorescence
 </p>
@@ Line 3,010: / Line 2,819: @@
 	</tr>
 </table></div>
-<!-- EDIT37 TABLE [38192-38898] -->
+<!-- EDIT37 TABLE [38837-39543] -->
 </div>
-<h4><a name="section100816" id="section100816">10/08/16</a></h4>
+<h4><a name="section10082016" id="section10082016">10/08/2016</a></h4>
 <div class="level4">
 <p>
-<br/>
+<strong>Fluorescence microscopy 33 (Nikon C2+ confocal microscope): Blocking-test A: spores + milk powder + GFP (in solution)</strong>
-<strong>Fluorescence microscopy 33 (Nikon C2+ confocal microscope): Blocking-test A: spores milk powder + GFP (in solution)</strong>
 </p>
 <p>
-–&gt;
+–&gt; spores and WT show similar levels of fluorescence
 </p>
 </div>
-<h4><a name="section101116" id="section101116">10/11/16</a></h4>
+<h4><a name="section10112016" id="section10112016">10/11/2016</a></h4>
 <div class="level4">
 <p>
-<br/>
+<strong>Sample preparation 34: Blocking-test B: spores + BSA + GFP (in solution)</strong>
-<strong>Sample preparation 34: Blocking-test B: BSA + GFP (in solution)</strong>
 </p>
 <ol>
@@ Line 3,078: / Line 2,883: @@
 	</tr>
 </table></div>
-<!-- EDIT38 TABLE [39474-39764] -->
 </div>
-<h4><a name="section101216" id="section101216">10/12/16</a></h4>
+<h4><a name="section10122016" id="section10122016">10/12/2016</a></h4>
 <div class="level4">
 <p>
-<br/>
+<strong>Fluorescence microscopy 34 (Nikon C2+ confocal microscope): Blocking-test B: spores + BSA + GFP (in solution)</strong>
 </p>
 <p>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:12.10.16_exp1_stamm9_100ms_60x.jpg" class="media" title="labprotocol:12.10.16_exp1_stamm9_100ms_60x.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:12.10.16_exp1_stamm9_100ms_60x.jpg" class="medialeft" align="left" alt="" width="200" /></a>
+–&gt; too little spores
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:12.10.16_exp1_stamm9_4s_60x.jpg" class="media" title="labprotocol:12.10.16_exp1_stamm9_4s_60x.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:12.10.16_exp1_stamm9_4s_60x.jpg" class="medialeft" align="left" alt="" width="200" /></a>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:12.10.16_exp_1_wt_mit_gfp_100ms_60x.jpg" class="media" title="labprotocol:12.10.16_exp_1_wt_mit_gfp_100ms_60x.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:12.10.16_exp_1_wt_mit_gfp_100ms_60x.jpg" class="medialeft" align="left" alt="" width="200" /></a>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:12.10.16_exp_1_wt_mit_gfp_100ms_60xa.jpg" class="media" title="labprotocol:12.10.16_exp_1_wt_mit_gfp_100ms_60xa.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:12.10.16_exp_1_wt_mit_gfp_100ms_60xa.jpg" class="medialeft" align="left" alt="" width="200" /></a>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:12.10.16_expstamm7.2_mit_gfp_100ms_60xb.jpg" class="media" title="labprotocol:12.10.16_expstamm7.2_mit_gfp_100ms_60xb.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:12.10.16_expstamm7.2_mit_gfp_100ms_60xb.jpg" class="medialeft" align="left" alt="" width="200" /></a>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:12.10.16_expstamm7.2_mit_gfp_4s_60xb.jpg" class="media" title="labprotocol:12.10.16_expstamm7.2_mit_gfp_4s_60xb.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:12.10.16_expstamm7.2_mit_gfp_4s_60xb.jpg" class="medialeft" align="left" alt="" width="200" /></a>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:12.10.16_exp_1_wt_ohne_gfp_100ms_60xa.jpg" class="media" title="labprotocol:12.10.16_exp_1_wt_ohne_gfp_100ms_60xa.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:12.10.16_exp_1_wt_ohne_gfp_100ms_60xa.jpg" class="medialeft" align="left" alt="" width="200" /></a>
-<a href="/igem2016/lib/exe/detail.php?id=labprotocol%3Amagic_bullets&amp;media=labprotocol:12.10.16_exp_1_wt_ohne_gfp_4s_60xa.jpg" class="media" title="labprotocol:12.10.16_exp_1_wt_ohne_gfp_4s_60xa.jpg"><img src="/igem2016/lib/exe/fetch.php?w=200&amp;media=labprotocol:12.10.16_exp_1_wt_ohne_gfp_4s_60xa.jpg" class="medialeft" align="left" alt="" width="200" /></a>
 </p>
 <div class="tags"><span>
@@ Line 3,107: / Line 2,904: @@
 </body>
-</html>
+</html> {{Freiburg/Footer}}
-{{Freiburg/Footer}}

Difference between revisions of "Team:Freiburg/NotebookTargeting"

Revision as of 16:15, 17 October 2016

Our Protocols