Team:EPF Lausanne/Microfluidics/Designing

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                <li><a href="https://2014.igem.org/Team:EPF_Lausanne/Microfluidics/Making/PartI">Part I</a></li>
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<h1 class="cntr">Microfluidics</h1>
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<img src="https://static.igem.org/mediawiki/2014/5/55/Chip_icon.png" alt="Chip icon" height="300" />
 
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<!--<p class="lead">Our Biopad is implemented in a microfluidic chip. This tool allows all kinds of analytical experiments and is increasingly used in biological research. From fabrication to applications, find out more about this awesome device here!</p>-->
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<a href="https://2014.igem.org/Team:EPF_Lausanne/Microfluidics" class="btn btn-primary pull-left" role="button">&lt;- Overview</a>
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<h3>Microfluidics and synthetic biology</h3>
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<a href="https://2014.igem.org/Team:EPF_Lausanne/Microfluidics/Making/PartI" class="btn btn-primary pull-right" role="button">Next step: Making a chip part I -&gt;</a>
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<p>Microfluidics is an efficient tool for biological experiments. Its fields of applications go from gene regulatory network analysis to antibody screening. Several laboratory techniques can be adapted to these devices, such as DNA amplification, protein separation or cell sorting.</p>  
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<h1 class="center">Designing a chip</h1>
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<p>The chips are generally fabricated from elastomeric materials, such as polydimethylsiloxane (PDMS) and contain micron-sized channels with integrated micromechanical tools (mixer, valve, pump…). This allows massive parallelisation as well as great modularity of the experiments.</p>
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<h2>The thought process of our designs</h2>
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<a href="https://static.igem.org/mediawiki/2014/7/71/Chip_diagram.png" data-lightbox="img1" data-title="The evolution of our designs"><img src="https://static.igem.org/mediawiki/2014/7/71/Chip_diagram.png" width="60%" class="pull-right img-right img-border"></a>
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<p>Most soluble reagents can be used, including DNA, proteins and small molecule libraries. As we focused our work on E.coli and S. cerevisiae, most of our experiments included culture of these species on-chip during our experiments. We first used the MITOMI chip which was invented in the lab of our supervisor Prof. Maerkl. We then designed new chips that were more adapted to stress the cells by pressure, as needed to implement the final “BioPad”.</p>
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<h3 id="smash">The SmashColi</h3>
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<p class="lead">When we started microfluidic experiments, the experiments required flowing cells in chambers and exposing them to different solutions and so we used the chip that was already available to us : the <a target="_blank" href="http://link.springer.com/protocol/10.1007%2F978-1-61779-292-2_6">MITOMI chip</a>.</p>
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<p class="lead">However for our project, where mechanical pressure will induce our touch sensitive bacteria, we thought of designing a new chip with bigger chambers (to increase the emission of the signal per chamber) and to add ‘huge buttons’ above the chambers to enable us to ‘squish the cells’, thus the SmashColi was designed.</p>
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<p>The major benefits of using microfluidic chips are:</p>
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<p class="lead">This chip is able to separate the array of chambers in 4, permitting us to flow in different cells and different solutions on one chip. Additionally, an input was designated for every 7 columns of buttons allowing us to put 4 different pressures on each row of cells. One row out of two was deprived of buttons to be used as a negative control.
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With all of this in mind, one chip is able to have 4 different cells and for each type of cells 4 pressures can be applied on them giving a total of 16 different experiments on just one chip!</p>
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<li>Low volume required (microliter range)</li>
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<li>High-throughput</li>
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<li>High precision and sensitive detection</li>
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<li>Cheap</li>
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<li>Wide range of applications</li>
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<li>Safe, enclosed environment (for more information go to the safety page)</li>
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<p class="lead">You may wonder why we didn’t just have chambers and press on the chip with a pen or our finger. Well the possibility of the SmashColi is that by applying the pressure through a machine, it is possible to quantify how the cells react towards a specific pressure. Once the cells were ready we would be able to quantify the intensity of the signal based on the pressure applied to them.</p>
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<p>Some examples of microfluidic experiments:</p>
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<a href="https://static.igem.org/mediawiki/2014/d/de/Microfdes1.png" data-lightbox="img1" data-title="Control layer of SmashColi"><img src="https://static.igem.org/mediawiki/2014/d/de/Microfdes1.png" width="46%"></a>
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<a href="https://static.igem.org/mediawiki/2014/f/f1/Microfdes2.png" data-lightbox="img1" data-title="Flow layer of SmashColi"><img src="https://static.igem.org/mediawiki/2014/f/f1/Microfdes2.png" width="48%"></a>
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<li>Transcription factors – DNA interactions</li>
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<li>Protein – protein interactions</li>
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<li>On-chip gene synthesis: protein expression from coding DNA</li>
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<li>On-chip chemostat chambers: can be used to trace the fate of a single bacterium or to grow bacteria/yeast</li>
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<li>Antibody characterisation</li>
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<h3>How does it work ?</h3>
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<p>Control layer design & Flow layer design.</p>
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<a href="https://static.igem.org/mediawiki/2014/0/0b/Microfdes5.png" data-lightbox="img1" data-title="Flow & control layers of SmashColi overlapped"><img src="https://static.igem.org/mediawiki/2014/0/0b/Microfdes5.png" class="img-responsive"></a>
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<p>Both layers overlapped.</p>
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<img src="https://static.igem.org/mediawiki/2014/0/03/Chip_sketch.png" alt="Chip sketch" class="cntr" width="100%" />
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<a href="https://static.igem.org/mediawiki/2014/0/07/BiopadFull.png" data-lightbox="img1" data-title="BioPad, final device design"><img src="https://static.igem.org/mediawiki/2014/0/07/BiopadFull.png" width="50%" class="pull-right img-right"></a>
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<h3 id="biopad_">Final device: the BioPad</h3>
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<p class="lead">For our final device, the idea was to have a large sized microfluidic chip where the cells can grow in chambers, each chambers will act as a ‘pixel’.</p>
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<p class="lead">The design is pretty simple, consisting of only a flow layer.</p>
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<ul>
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<li>a. Disassembled view of a microfluidic chip showing all the different components and the region where bacteria/yeasts are located</li>
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<li>b. Cross section of the chip showing how a valve works: when pressure is applied in the control channel, the ceiling of the flow layer is pushed against the glass slide, which closes the flow channel</li>
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<li>c. When pressure is retrieved, the ceiling elevates again, which opens the flow channel</li>
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<p>A standard microfluidic chip is a grid of interconnected channels and chambers. It is usually composed of one or two PDMS layers placed on a glass slide. In our case we used two layers, the so called flow layer and control layer. The bacteria are enclosed between the flow layer and the glass slide. By its shape, the flow layer is responsible for the patterns of the chip. In our case, the pattern consists of several parallel rows of chambers. The control layer comes on top of the flow layer and allows to open or close valves by pressing or releasing water in the corresponding channels. Thus a mechanical pressure can be applied from the control layer on the flow layer, enabling a precise compartmentalization of the chip.</p>
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<h3 id="filter">The FilterColi</h3>
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<br/>
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<p class="lead">One of the experiments that we did at the end of the summer was to expose the cells to different solutions by flowing it in and diffuse it in the chambers. However we saw that it wasn’t very effective and so we designed a new chip were the solution is flowed through the cell and so the cells are directly in contact with the solution. </p>
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<p class="lead">The FilterColi was the answer to our problem. By adding filters in the chambers and another flow channel below the chambers, we would be able to flow solution through the cells. </p>
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<p>Once the chip is ready to be used, small tubings of 0.35mm diameter are plugged in the inlets of the chip (see gif below). The tubings that are plugged in the control inlets are loaded with water and enable the opening or closing of valves. The tubings that are plugged into the flow inlets are used to flow bacteria/yeast or various solutions in the chambers. </p>
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<a href="https://static.igem.org/mediawiki/2014/8/89/FilterControl.png" data-lightbox="img1" data-title="Control layer of FilterColi"><img src="https://static.igem.org/mediawiki/2014/d/de/Microfdes1.png" width="46%"></a>
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<a href="https://static.igem.org/mediawiki/2014/4/48/FilterFlow.png" data-lightbox="img1" data-title="Flow layer of FilterColi"><img src="https://static.igem.org/mediawiki/2014/f/f1/Microfdes2.png" width="48%"></a>
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<p>Control layer design & Flow layer design.</p>
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<a href="https://static.igem.org/mediawiki/2014/c/c8/Filterfull.png" data-lightbox="img1" data-title="Flow & control layers of FilterColi overlapped"><img src="https://static.igem.org/mediawiki/2014/c/c8/Filterfull.png" class="img-responsive"></a>
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<p>Both layers overlapped.</p>
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<p>Picture of the MITOMI Chip and our Smash-Coli chip</p>
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<h3 id="clean">The CleanColi</h3>
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<p class="lead">As we did the safety issues for Microfluidics, we brainstormed to make an “on chip waste treatment”. The idea was to implement mechanisms in the chip, which will treat the cells that were to exit the array of chambers.<br/>
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<div class="cntr">
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<a href="https://static.igem.org/mediawiki/2014/f/ff/Clean.png" data-lightbox="img1" data-title="CleanColi device"><img src="https://static.igem.org/mediawiki/2014/f/ff/Clean.png" width="80%"></a><br/><br/>
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The process we created is the following : <br/><br/><br/><br/>
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<a href="https://static.igem.org/mediawiki/2014/0/05/Serpentine.png" data-lightbox="img1" data-title="Serpentine to mix the Bleach or lysis in the flow"><img src="https://static.igem.org/mediawiki/2014/0/05/Serpentine.png" width="48%"></a><br/>
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    <div class="thumbnail">
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<a href="https://static.igem.org/mediawiki/2014/c/c6/Mitomi_che.PNG" data-lightbox="image-1" data-title="Mitomi"><img src="https://static.igem.org/mediawiki/2014/c/c6/Mitomi_che.PNG" alt="Mitomi" width="200" /></a>
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        <p>MITOMI chip filled with bacteria expressing GFP</p>
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<a href="https://static.igem.org/mediawiki/2014/5/51/Killcoli.PNG" data-lightbox="image-1" data-title="Smash-coli"><img src="https://static.igem.org/mediawiki/2014/5/51/Killcoli.PNG" alt="Killcoli" width="200" /></a>
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        <p>“Smash-coli” chip, here with expression of RFP</p>
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Step 1. The cells leave the array of chambers and are flowed in a serpentine circuit with lysis buffer to mix the cells and the lysis.<br/>
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Step 2. The waste is then flowed in another serpentine circuit with bleach.<br/><br/><br/><br/>
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<a href="https://static.igem.org/mediawiki/2014/3/35/Filter.png" data-lightbox="img1" data-title="A filter chamber to block all the cells and debris"><img src="https://static.igem.org/mediawiki/2014/3/35/Filter.png" width="48%"></a><br/>
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Step 3. The waste is retained in a last big chamber where they are confronted to smaller and smaller filters to keep the debris in the chamber and only liquid will be flowed out of the chip.<br/><br/><br/><br/>
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<a href="https://static.igem.org/mediawiki/2014/9/93/Heater.png" data-lightbox="img1" data-title="A microheater to denature the cells or proteins that survived the rest of the process"><img src="https://static.igem.org/mediawiki/2014/9/93/Heater.png" width="48%"></a><br/>
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Step 4. The cells enter a big chamber located above a microheater inducing a local temperature of 95°C.<br/>
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  <li><a href="https://2014.igem.org/Team:EPF_Lausanne/Microfluidics/Designing">Designing a chip</a></li>
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<a href="https://2014.igem.org/Team:EPF_Lausanne/Microfluidics/Making/PartI" class="btn btn-primary pull-right" role="button">Next step: Making a chip part I -&gt;</a>
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                Making a chip <b class="caret"></b>
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                <li><a href="https://2014.igem.org/Team:EPF_Lausanne/Microfluidics/Making/PartI">Part I</a></li>
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                <li><a href="https://2014.igem.org/Team:EPF_Lausanne/Microfluidics/Making/PartII">Part II</a></li>
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        <!--<li class="active"><a href="#syntheticbiology">Microfluidics & synthetic biology</a></li>-->
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        <li><a href="#smash">SmashColi</a></li>
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        <li><a href="#biopad_">BioPad</a></li>
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        <li><a href="#filter">FilterColi</a></li>
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Latest revision as of 01:35, 18 October 2014

<- Overview Next step: Making a chip part I ->

Designing a chip



The thought process of our designs


The SmashColi


When we started microfluidic experiments, the experiments required flowing cells in chambers and exposing them to different solutions and so we used the chip that was already available to us : the MITOMI chip.

However for our project, where mechanical pressure will induce our touch sensitive bacteria, we thought of designing a new chip with bigger chambers (to increase the emission of the signal per chamber) and to add ‘huge buttons’ above the chambers to enable us to ‘squish the cells’, thus the SmashColi was designed.

This chip is able to separate the array of chambers in 4, permitting us to flow in different cells and different solutions on one chip. Additionally, an input was designated for every 7 columns of buttons allowing us to put 4 different pressures on each row of cells. One row out of two was deprived of buttons to be used as a negative control. With all of this in mind, one chip is able to have 4 different cells and for each type of cells 4 pressures can be applied on them giving a total of 16 different experiments on just one chip!

You may wonder why we didn’t just have chambers and press on the chip with a pen or our finger. Well the possibility of the SmashColi is that by applying the pressure through a machine, it is possible to quantify how the cells react towards a specific pressure. Once the cells were ready we would be able to quantify the intensity of the signal based on the pressure applied to them.

Control layer design & Flow layer design.

Both layers overlapped.


Final device: the BioPad


For our final device, the idea was to have a large sized microfluidic chip where the cells can grow in chambers, each chambers will act as a ‘pixel’.

The design is pretty simple, consisting of only a flow layer.

The FilterColi


One of the experiments that we did at the end of the summer was to expose the cells to different solutions by flowing it in and diffuse it in the chambers. However we saw that it wasn’t very effective and so we designed a new chip were the solution is flowed through the cell and so the cells are directly in contact with the solution.

The FilterColi was the answer to our problem. By adding filters in the chambers and another flow channel below the chambers, we would be able to flow solution through the cells.

Control layer design & Flow layer design.

Both layers overlapped.




The CleanColi


As we did the safety issues for Microfluidics, we brainstormed to make an “on chip waste treatment”. The idea was to implement mechanisms in the chip, which will treat the cells that were to exit the array of chambers.



The process we created is the following :




Step 1. The cells leave the array of chambers and are flowed in a serpentine circuit with lysis buffer to mix the cells and the lysis.
Step 2. The waste is then flowed in another serpentine circuit with bleach.




Step 3. The waste is retained in a last big chamber where they are confronted to smaller and smaller filters to keep the debris in the chamber and only liquid will be flowed out of the chip.




Step 4. The cells enter a big chamber located above a microheater inducing a local temperature of 95°C.

<- Overview Next step: Making a chip part I ->

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