Difference between revisions of "Team:Technion Israel/Hardware"

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        <title>S.tar, by iGEM Technion 2016</title>
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<h3>★  ALERT! </h3>
 
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<p>This page is used by the judges to evaluate your team for the<a href="https://2016.igem.org/Judging/Awards"> Best Hardware award</a>. </p>
 
  
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<p>iGEM is about making teams of students making synthetic biology projects. We encourage teams to work with parts and build biological devices in the lab. But we are inclusive and want all teams to work on many other types of problems in synbio. Robotic assembly, microfluidics, low cost equipment and measurement hardware are all areas ripe for innovation in synbio. </p>
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.ocenter .img-responsive {
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Teams who are interested in working with hardware as a side project are encouraged to apply for the hardware award.  
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<img src="https://static.igem.org/mediawiki/2016/4/44/T--Technion_Israel--description.jpg" class="img-responsive img-center cont_cover" width="100%">
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<h1 class="text-center"><u>Hardware</u></h1>
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<div class="col-md-12 col-sm-12">
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<h2>FlashLab</h2>
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</div>
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<br>
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<br>
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<p class="text-justify">
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We developed our first FlashLab prototypes around on the commercial fluidic chip
 +
(IBIDI sticky–Slide I Luer 0.8). The commercial chip is designed mostly for performing
 +
cell culture experiments under shear stress, with custom specific bottom.
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</p>
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</div><!--
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--><div class="col-md-6 col-sm-12 vcenter"><!--6 img div-->
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<a class="pop ocenter">
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<img src="https://static.igem.org/mediawiki/2016/9/91/T--Technion_Israel--Hardware1.jpg" class="img-responsive img-center" width="450" style="cursor: pointer;">
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</a>
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<p class="text-center"><b>Fig. 1:</b> The geometry of the commercial fluidic chip</p>
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</div>
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</div>
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<br>
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<br>
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<br>
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<p class="text-center"><b>Table 1:</b> <b>Here there should be a table</b></p>
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<img src="" class="img-responsive img-center" width="450" style="cursor: pointer;">
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</a>
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</div>
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<br>
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<br>
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<div class="row">
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<div class="col-md-12 col-sm-12">
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<p class="text-justify">The chip is made out of plastic and the bottom can be closed by sticking it to a glass slide.</B></p>
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<a class="pop ocenter">
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<img src="https://static.igem.org/mediawiki/2016/d/d0/T--Technion_Israel--Hardware2.png" class="img-responsive img-center" width="450" style="cursor: pointer;">
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</a>
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<p class="text-center"><b>Fig. 2:</b> Setup of commercial fluidic chip.</p>
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<br>
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<br>
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<p class="text-justify">
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The commercial chip worked for preliminary testing but not idle for our uses: The entry
 +
slots are relatively wide, making it difficult to load the sample in a uniform and
 +
even fashion. This affects the diffusion of the chemo-repellent in the channel and
 +
reduces the overall accuracy of the device. Also, the channel is relatively shallow,
 +
forcing the use of high concentration of bacteria to get a visible signal. Storing a
 +
large amount of bacteria in a confined area might cause oxygen shortage that will
 +
harm bacterial motility.<br>
 +
<br>
 +
We designed a new chip that confronts those problems:<br>
 +
<b>-</b> Reducing the radius of the entry slot will enable a controlled insertion of the
 +
sample. The smaller slot will slow down any flow (for example, flow caused by loading
 +
sample from a syringe). Also, this will fix the diffusion source at permanent place
 +
for all of our experiments.<br>
 +
<b>-</b> Shaping the channel as funnel will cause bacteria to concentrate as they
 +
move away from chemo-repellents (from left to right).<br>
 +
<b>-</b> A deeper channel will enable to see more color in the same bacterial
 +
concentration than in the commercial chip, while reducing the risk of lack of oxygen.<br>
 +
<br>
 +
</p>
 +
</div>
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</div>
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<br>
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<br>
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<br>
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<a class="pop ocenter">
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<img src="https://static.igem.org/mediawiki/2016/3/38/T--Technion_Israel--Hardware3.png" class="img-responsive img-center" width="450" style="cursor: pointer;">
 +
</a>
 +
<p class="text-center"><b>Fig. 3:</b> The geometry of designed fluidic chip.</p>
 +
</div>
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</div>
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<br>
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<br>
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<br>
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<p class="text-justify">
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The new chip was fabricated in two methods: PDMS chip and Dolomite 3D printer.
 +
</p>
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</div>
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</div>
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<div class="col-md-12 col-sm-12">
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<h2>PDMS</h2>
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</div>
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</div>
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<div class="row">
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<div class="col-md-12 col-sm-12">
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<p class="text-justify">
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PDMS is the considered the standard for microfluidic fabrication in labs.
 +
PDMS is optically clear, and, in general, inert, non-toxic, and non-flammable.<br>
 +
<br>
 +
Overview on the fabrication process:<br>
 +
<b>1. </b>Designed a two parts mold using SolidWorks software: cover and base.<br>
 +
<b>2. </b>Printing the mold using Ultimaker 2 Etentended+ 3D printer.<br>
 +
<b>3. </b>Mixing the polymer base and curing agent at 10:1 weight ratio (with the advantage to the polymer base). fill the mold with the mix.<br>
 +
<b>4. </b>Put the mold inside desiccator to degas for 2 hours.<br>
 +
<b>5. </b>Insert the mold inside the oven at 70 C for 3 hours.<br>
 +
<b>6. </b>carefully take off the mold’s cover and then cut out the PDMS chip.<br>
 +
<b>7. </b>Stick the PDMS chip into thin cover glass (0.3 mm) using silicon glue.*<br>
 +
*Whan making a PDMS chip.<br>
 +
</p>
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</div>
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<div class="row">
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<a class="pop ocenter">
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<p class="text-center"><b>Table 2:</b> <b>Here there should be a table</b></p>
 +
<img src="" class="img-responsive img-center" width="450" style="cursor: pointer;">
 +
</a>
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</div>
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</div>
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<div class="row">
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<div class="col-md-12 col-sm-12">
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<p class="text-justify">
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The foollowing scheam describes the mentioned process:
 +
</p>
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</div>
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</div>
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 +
<br>
 +
<br>
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<br>
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<div class="row">
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<div class="col-md-12 col-sm-12">
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<a class="pop ocenter">
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<img src="https://static.igem.org/mediawiki/2016/5/52/T--Technion_Israel--Hardware4.jpg" class="img-responsive img-center" width="500" style="cursor: pointer;">
 +
</a>
 +
<p class="text-center"><b>Fig. 4:</b> PDMS chip fabrication process</p>
 +
</div>
 +
</div>
 +
 
 +
<br>
 +
<br>
 +
<br>
 +
 
 +
<div class="row">
 +
<div class="col-md-12 col-sm-12">
 +
<p class="text-justify">
 +
<b>Designing the mold:</b><br>
 +
The mold is composed out of two parts to create unique geometric and for
 +
easier extraction of the PDMS out of the mold.<br>
 +
</p>
 +
</div>
 +
</div>
 +
 
 +
<!--6 text - 6 img div-->
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<div class="row">
 +
<div class="col-md-6 col-sm-12 vcenter"><!--6 text-->
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<p class="text-justify">
 +
<u>The base</u><br>
 +
<b>-</b> The cone on the base of the floor is make the funnel shape of the chip ((a) in figure 2).<br>
 +
<b>-</b> Small slits were made in the walls of the base to position the cover accurately.<br>
 +
<b>-</b> The overall size was determent so the chip will fit on standard microscope cover
 +
slide. this will enable us to run experiments under a microscope easily.<br>
 +
</p>
 +
</div><!--
 +
--><div class="col-md-6 col-sm-12 vcenter"><!--6 img div-->
 +
<a class="pop ocenter">
 +
<img src="https://static.igem.org/mediawiki/2016/4/48/T--Technion_Israel--Hardware5.jpg" class="img-responsive img-center" width="450" style="cursor: pointer;">
 +
</a>
 +
<p class="text-center"><b>Fig. 5:</b> The geometry of the mold</p>
 +
</div>
 +
</div>
 +
 
 +
<br>
 +
<br>
 +
<br>
 +
 
 +
<!--6 text - 6 img div-->
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<div class="row">
 +
<div class="col-md-6 col-sm-12 vcenter"><!--6 text-->
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<p class="text-justify">
 +
<u>The cover</u><br>
 +
<b>-</b> Four rods coming out of the sides of the cover for easy extraction of the cover
 +
when taking out the PDMS.<br>
 +
<b>-</b> The ramp is to insure that the channel will be inserted inside the PDMS and
 +
getting the wanted channel height.<br>
 +
<b>-</b> The cover is made smaller than the base for a good fit and for letting out
 +
any gas that might have been caught when inserting it. Those gases, if
 +
left in will expend in the oven and cause deformation in the chip. <br>
 +
</p>
 +
</div><!--
 +
--><div class="col-md-6 col-sm-12 vcenter"><!--6 img div-->
 +
<a class="pop ocenter">
 +
<img src="https://static.igem.org/mediawiki/2016/f/fd/T--Technion_Israel--Hardware6.jpg" class="img-responsive img-center" width="450" style="cursor: pointer;">
 +
</a>
 +
<p class="text-center"><b>Fig. 6:</b> The geometry of the cover.</p>
 +
</div>
 +
</div>
 +
 
 +
<br>
 +
<br>
 +
<br>
 +
 
 +
<!-- 12 text div -->
 +
<div class="row">
 +
<div class="col-md-12 col-sm-12">
 +
<p class="text-justify">
 +
Printing the mold using Ultimaker 2 Etentended+. This 3d printer was chosen
 +
because of its high accuracy (X,Y,Z =12.5, 12.5, 5 micron) and due that the
 +
fact that the polymer it uses (PLA) can be heated to relatively high temperatures
 +
without changing form (TG=60-65 C) and does not reacts to the PDMS. Another
 +
benefits of 3D printing are the low price and fast manufacturing time: We
 +
printed our mold for about 25$, and it took about 6 hours. 
 +
</p>
 +
</div>
 +
</div>
 +
 
 +
<br>
 +
<br>
 +
<br>
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 +
<!-- Mini headline -->
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<div class="row">
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<div class="col-md-12 col-sm-12">
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<h2>Dolomite Fluidic Factory</h2>
 +
</div>
 +
</div>
 +
 
 +
<!--6 text - 6 img div-->
 +
<div class="row">
 +
<div class="col-md-6 col-sm-12 vcenter"><!--6 text-->
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<p class="text-justify">
 +
Fluidic Factory enables fast prototyping of microfluidic chips, manifolds and
 +
connectors using COC (FDA approved, biocompatible, translucent and robust polymer).
 +
Printing the chip toke about 3 hours and was made straight from computer model.
 +
This technology just came out this year and we are the first iGEM group to ever use it.
 +
</p>
 +
</div><!--
 +
--><div class="col-md-6 col-sm-12 vcenter"><!--6 img div-->
 +
<a class="pop ocenter">
 +
<img src="https://static.igem.org/mediawiki/2016/5/5c/T--Technion_Israel--Hardware7.jpg" class="img-responsive img-center" width="450" style="cursor: pointer;">
 +
</a>
 +
<p class="text-center"><b>Fig. 7:</b> Factory chip fabrication process.</p>
 +
</div>
 +
</div>
 +
 
 +
<br>
 +
 
 +
<!-- 12 text div -->
 +
<div class="row">
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<div class="col-md-12 col-sm-12">
 +
<p class="text-justify">
 +
<b>Results:</b> We had some difficulties preventing the channels from collapsing in the "Dolomite" printer.
 +
While not achieving a usable chip, we believe that this technology shows a lot of promise.<br>
 +
</p>
 +
</div>
 +
</div>
 +
 
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<br>
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<br>
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<br>
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 +
<!-- Mini headline -->
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<div class="row">
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<div class="col-md-12 col-sm-12">
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<h2>Quantitative test for bacterial concentration</h2>
 +
</div>
 +
</div>
 +
 
 +
<!--6 text - 6 img div-->
 +
<div class="row">
 +
<div class="col-md-6 col-sm-12 vcenter"><!--6 text-->
 +
<p class="text-justify">
 +
As the bacteria sense either the repellent or the attractant in a certain direction creating
 +
a cluster. Our bacteria may be visible, but at low concentrations the cluster might be
 +
difficult to distinguish. Through the bacterial concentration test, it will be possible
 +
to determine quantitatively whether a cluster was formed or not.<br>
 +
The system is composed of two independent electrical circuits as illustrated in.
 +
</p>
 +
</div><!--
 +
--><div class="col-md-6 col-sm-12 vcenter"><!--6 img div-->
 +
<a class="pop ocenter">
 +
<img src="https://static.igem.org/mediawiki/2016/7/78/T--Technion_Israel--Hardware8.jpg" class="img-responsive img-center" style="cursor: pointer;">
 +
</a>
 +
<p class="text-center"><b>Fig. 8:</b>  The electricl circuits. (a) contains a LED a voltage source of 5v,
 +
(b) contains a photoresistor,  a resistor and a voltage source.</p>
 +
</div>
 +
</div>
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<br>
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<br>
 +
<br>
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<!-- 12 text div -->
 +
<div class="row">
 +
<div class="col-md-12 col-sm-12">
 +
<p class="text-justify">
 +
<b>Principle of Operation:</b><br>
 +
The LED, the chip and the photoresistor are placed in a dark box to prevent undesired light
 +
leakages and reflections . The LED emits light at 585-595nm (the specs are in the appendix),
 +
which passes through the chip and partially absorbed by the bacteria. The transmitted light
 +
reaches to the photoresistor  and causes a decrease in its resistance. As a result, the
 +
voltage that falls on the photoresistor decreases and similarly the voltage that falls
 +
on the resistor  increases. Then, the signal produced by the photoresistor is converted
 +
to a digital signal that goes to the computer. Finally, the output  voltage is displayed
 +
with a GUI user-friendly interface as shown in Fig 2.<br>
 +
</p>
 +
</div>
 +
</div>
 +
 
 +
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 +
<br>
 +
<br>
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<img src="https://static.igem.org/mediawiki/2016/e/ec/T--Technion_Israel--Hardware9.jpg" class="img-responsive img-center" style="cursor: pointer;">
 +
</a>
 +
<p class="text-center"><b>Fig. 9:</b> GUI-interface.</p>
 +
</div>
 +
</div>
 +
 
 +
<br>
 +
<br>
 +
<br>
 +
 
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<div class="col-md-12 col-sm-12">
 +
<p class="text-justify">
 +
The left side of the interface is used for testing the system and the right side serves
 +
for taking measurments. The user can change some parameters by the GUI interface like the
 +
number of samples that the system collects and at which frequency.<br>
 +
GUI displays a graph of the voltage that falls on the resistor as a function of number
 +
of samples, which is equivalent to time.<br>
 +
After taking reference, the user can take a measurement. The System reading is the ratio
 +
between the average of the sample voltage taken in the measurement divided by the samples
 +
voltage average of the reference.<br>
 +
</p>
 +
</div>
 +
</div>
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<br>
 +
<br>
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<br>
 +
 
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<!-- 12 text div -->
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<div class="row">
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<div class="col-md-12 col-sm-12">
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<p class="text-justify">
 +
<b>GUI:</b>
 +
Gui is a graphical user interface  is a type of user interface that allows users to interact
 +
with electronic devices through graphical icons and visual indicators such as secondary notation,
 +
instead of text-based user interfaces, typed command labels or text navigation.<br>
 +
In order to use GUI, it is required to download Arduino I/O toolbox.<br>
 +
In our project we used matlab as programming language. The matlab code is in the appendix section.<br>
 +
</p>
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</div>
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<b>Deriving the relationship between the voltage that falls on the resistor and the O.D of the bacterial solution:
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According to the voltage divider rule, the voltage that falls on the resistor
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in the photoresistor circuit (VR) is equal to
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According to “Emant”, the relationship between the resistance RL of a typical LDR and the light intensity is:
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Where LUX is the light intensity that reaches the photoresistor.br
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<br>
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<br>
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Combining  Equation 1 and Equation 2:
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By definition:
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Where I<sub>0</sub> is the light intensity emitted from the LED and A is the optical
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density of the bacterial concentration inside the chip.<br>
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<br>
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From Equation 4 it can be derived that VR decreases as A increases.<br>
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<b>Bill of materials</b>
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<p class="text-center"><b>Table ###:</b> <b>Here there should be a table</b></p>
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<img src="" class="img-responsive img-center" width="450" style="cursor: pointer;">
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For the matlab code, see: <b>!@#$%^&^%$</b>
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1.  Calloway, D. (1997). Beer-Lambert Law. Journal of Chemical Education, 74(7), 744. http://doi.org/10.1021/ed074p744.3<br>
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<br>
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2. XYZ<br>
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<br>
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3. XYZ <br>
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<br>
 
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<h5>Inspiration</h5>
 
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<p>You can look at what other teams did to get some inspiration! <br />
 
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Here are a few examples:</p>
 
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<ul>
 
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<li><a href="https://2015.igem.org/Team:TU_Delft">2015 TU Delft  </a></li>
 
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<li><a href="https://2015.igem.org/Team:TU_Darmstadt">2015 TU Darmstadt</a></li>
 
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<li><a href="https://2015.igem.org/Team:Cambridge-JIC">2015 Cambridge JIC</a></li>
 
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</ul>
 
  
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Revision as of 00:55, 15 October 2016

S.tar, by iGEM Technion 2016

S.tar, by iGEM Technion 2016

Hardware

FlashLab



We developed our first FlashLab prototypes around on the commercial fluidic chip (IBIDI sticky–Slide I Luer 0.8). The commercial chip is designed mostly for performing cell culture experiments under shear stress, with custom specific bottom.

Fig. 1: The geometry of the commercial fluidic chip




Table 1: Here there should be a table



The chip is made out of plastic and the bottom can be closed by sticking it to a glass slide.

Fig. 2: Setup of commercial fluidic chip.




The commercial chip worked for preliminary testing but not idle for our uses: The entry slots are relatively wide, making it difficult to load the sample in a uniform and even fashion. This affects the diffusion of the chemo-repellent in the channel and reduces the overall accuracy of the device. Also, the channel is relatively shallow, forcing the use of high concentration of bacteria to get a visible signal. Storing a large amount of bacteria in a confined area might cause oxygen shortage that will harm bacterial motility.

We designed a new chip that confronts those problems:
- Reducing the radius of the entry slot will enable a controlled insertion of the sample. The smaller slot will slow down any flow (for example, flow caused by loading sample from a syringe). Also, this will fix the diffusion source at permanent place for all of our experiments.
- Shaping the channel as funnel will cause bacteria to concentrate as they move away from chemo-repellents (from left to right).
- A deeper channel will enable to see more color in the same bacterial concentration than in the commercial chip, while reducing the risk of lack of oxygen.




Fig. 3: The geometry of designed fluidic chip.




The new chip was fabricated in two methods: PDMS chip and Dolomite 3D printer.

PDMS

PDMS is the considered the standard for microfluidic fabrication in labs. PDMS is optically clear, and, in general, inert, non-toxic, and non-flammable.

Overview on the fabrication process:
1. Designed a two parts mold using SolidWorks software: cover and base.
2. Printing the mold using Ultimaker 2 Etentended+ 3D printer.
3. Mixing the polymer base and curing agent at 10:1 weight ratio (with the advantage to the polymer base). fill the mold with the mix.
4. Put the mold inside desiccator to degas for 2 hours.
5. Insert the mold inside the oven at 70 C for 3 hours.
6. carefully take off the mold’s cover and then cut out the PDMS chip.
7. Stick the PDMS chip into thin cover glass (0.3 mm) using silicon glue.*
*Whan making a PDMS chip.

Table 2: Here there should be a table

The foollowing scheam describes the mentioned process:




Fig. 4: PDMS chip fabrication process




Designing the mold:
The mold is composed out of two parts to create unique geometric and for easier extraction of the PDMS out of the mold.

The base
- The cone on the base of the floor is make the funnel shape of the chip ((a) in figure 2).
- Small slits were made in the walls of the base to position the cover accurately.
- The overall size was determent so the chip will fit on standard microscope cover slide. this will enable us to run experiments under a microscope easily.

Fig. 5: The geometry of the mold




The cover
- Four rods coming out of the sides of the cover for easy extraction of the cover when taking out the PDMS.
- The ramp is to insure that the channel will be inserted inside the PDMS and getting the wanted channel height.
- The cover is made smaller than the base for a good fit and for letting out any gas that might have been caught when inserting it. Those gases, if left in will expend in the oven and cause deformation in the chip.

Fig. 6: The geometry of the cover.




Printing the mold using Ultimaker 2 Etentended+. This 3d printer was chosen because of its high accuracy (X,Y,Z =12.5, 12.5, 5 micron) and due that the fact that the polymer it uses (PLA) can be heated to relatively high temperatures without changing form (TG=60-65 C) and does not reacts to the PDMS. Another benefits of 3D printing are the low price and fast manufacturing time: We printed our mold for about 25$, and it took about 6 hours.




Dolomite Fluidic Factory

Fluidic Factory enables fast prototyping of microfluidic chips, manifolds and connectors using COC (FDA approved, biocompatible, translucent and robust polymer). Printing the chip toke about 3 hours and was made straight from computer model. This technology just came out this year and we are the first iGEM group to ever use it.

Fig. 7: Factory chip fabrication process.


Results: We had some difficulties preventing the channels from collapsing in the "Dolomite" printer. While not achieving a usable chip, we believe that this technology shows a lot of promise.




Quantitative test for bacterial concentration

As the bacteria sense either the repellent or the attractant in a certain direction creating a cluster. Our bacteria may be visible, but at low concentrations the cluster might be difficult to distinguish. Through the bacterial concentration test, it will be possible to determine quantitatively whether a cluster was formed or not.
The system is composed of two independent electrical circuits as illustrated in.

Fig. 8: The electricl circuits. (a) contains a LED a voltage source of 5v, (b) contains a photoresistor, a resistor and a voltage source.




Principle of Operation:
The LED, the chip and the photoresistor are placed in a dark box to prevent undesired light leakages and reflections . The LED emits light at 585-595nm (the specs are in the appendix), which passes through the chip and partially absorbed by the bacteria. The transmitted light reaches to the photoresistor and causes a decrease in its resistance. As a result, the voltage that falls on the photoresistor decreases and similarly the voltage that falls on the resistor increases. Then, the signal produced by the photoresistor is converted to a digital signal that goes to the computer. Finally, the output voltage is displayed with a GUI user-friendly interface as shown in Fig 2.




Fig. 9: GUI-interface.




The left side of the interface is used for testing the system and the right side serves for taking measurments. The user can change some parameters by the GUI interface like the number of samples that the system collects and at which frequency.
GUI displays a graph of the voltage that falls on the resistor as a function of number of samples, which is equivalent to time.
After taking reference, the user can take a measurement. The System reading is the ratio between the average of the sample voltage taken in the measurement divided by the samples voltage average of the reference.




GUI: Gui is a graphical user interface is a type of user interface that allows users to interact with electronic devices through graphical icons and visual indicators such as secondary notation, instead of text-based user interfaces, typed command labels or text navigation.
In order to use GUI, it is required to download Arduino I/O toolbox.
In our project we used matlab as programming language. The matlab code is in the appendix section.




Deriving the relationship between the voltage that falls on the resistor and the O.D of the bacterial solution: According to the voltage divider rule, the voltage that falls on the resistor in the photoresistor circuit (VR) is equal to

According to “Emant”, the relationship between the resistance RL of a typical LDR and the light intensity is:

Where LUX is the light intensity that reaches the photoresistor.br

Combining Equation 1 and Equation 2:

By definition:


Where I0 is the light intensity emitted from the LED and A is the optical density of the bacterial concentration inside the chip.

From Equation 4 it can be derived that VR decreases as A increases.

Bill of materials

Table ###: Here there should be a table



For the matlab code, see: !@#$%^&^%$

Referances

1. Calloway, D. (1997). Beer-Lambert Law. Journal of Chemical Education, 74(7), 744. http://doi.org/10.1021/ed074p744.3

2. XYZ

3. XYZ




S.tar, by iGEM Technion 2016