Difference between revisions of "Team:Munich/Hardware"

 
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<tr id="hardwareFrontPage">
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<font size=7 color=#51a7f9><b style="color: #51a7f9">Hardware</b></font>
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<table width=320>
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<tr>
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<td  colspan = 6 align="left">
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<p class="introduction">
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Our pathogen detection approach relies on Cas13a digesting RNA. A common way of monitoring RNase activityis using commercially available RNaseAlert consisting of a fluorescent RNA beacon. This is impractical for in-fieldapplications because commercial fluorescence detectors are expensive and inconveniently large. We therefore makeour pathogen detection system fit for in-field applications by developing a cheap and handy fluorescence detector. Al-though many previous iGEM teams constructed fluorescence detectors, we could not find any that had a high enoughsensitivity or the ability to measure fluorescence quantitatively. We therefore constructed a detector matching ourrequirements and compared it to others in a cost vs sensitivity diagram.
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                </p>
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<p class="introduction">
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Our detector is paper-based and can detect fluorescein concentrations down to 200 nM. The detector is able to automatically
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measure fluorescence in units of equivalent fluorescein concentrations. It fits in a pipette box and costs less
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than 15 $. We were able to measure a time trace of Cas13a digesting RNaseAlert with our detector. For comparison
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we also measured a positive control containing RNase A and a negative control containing only RNaseAlert. The
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data are displayed in the figure bellow.
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                </p>
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<p class="introduction">
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The time traces show an enzymatic reaction taking place on filter paper. This proves that our detector is sensitive
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enough and meets our requirements. However the detector is not limited to our specific application but can be used
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for the detection of any fluorescence signal in biological or chemical systems. We therefore think that our detector
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can benefit other iGEM teams and research groups that want to make fluorescence based detection fit for in-field
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applications.
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                </p>
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<a href="/Team:Munich/Hardware/QuakeValve"><img class="picture1" src="https://static.igem.org/mediawiki/2017/0/08/T--Munich--Overlay.png"></a>
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<h3>Overall Design</h3>
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<p> 
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Light from a blue LED is filtered by a blue filter foil and excites fluorophores on a filter paper. The excitation light
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is blocked by an orange filter foil while the emission light from the fluoroscopes passes through the orange filter foil
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and illuminates a light dependent resistor (LDR). The LDR changes its resistance corresponding to the intensity
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of the fluorescence light.Finally an Arduino Nano measures the resistance via a voltage divider and calculates the
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fluorophore concentration. The two figures bellow show this overall design and the operational detector.</p>
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<a href="/Team:Munich/Hardware/SampleProcessing"><img id="picture3" src="https://static.igem.org/mediawiki/2017/0/08/T--Munich--Overlay.png"></a>
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<h3>Components</h3></td></tr>
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<a href="/Team:Munich/Hardware/Paperstrip"><img id="picture2" src="https://static.igem.org/mediawiki/2017/0/08/T--Munich--Overlay.png"></a>
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<h4>Micro Controller</h4>
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<p> 
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<a href="/Team:Munich/Hardware/Detector"><img id="picture4" src="https://static.igem.org/mediawiki/2017/0/08/T--Munich--Overlay.png"></a>
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We used an Arduino Nano for automatized data collection. This micro controller has analog pins that can measure
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voltages from 0 to 5 V and gives an integer from 0 to 1023 as output. The micro controller is connected via an USB port with a computer or smart-phone where the data can be processed further.</p>
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<h4>Light dependent resistor (LDR)</h4>
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<p id="equation1"> 
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<div class="captionPicture">
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For the detection of fluorescence light we used a light depending resistor (LDR). A LDR decreases its resistance <i>R<sub>LDR</sub></i> with increasing light intensity <i>I</i>. The dependence of the resistance <i>R<sub>LDR</sub></i> on the light intensity <i>I</i> is</p>
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<div class="equationDiv"><img src="https://static.igem.org/mediawiki/2017/3/33/T--Munich--Hardware_equation1.png"><span>(1)</span></div>
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<p>
 
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where γ is a parameter depending on the type of resistor being used and can even differ for LDRs with the same type
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Complete overview of all modular hardware parts in our pathogen detection system. Shown are counterclockwise and starting in in the upper left corner: The Quake valve that controls fluid flow, our sample processing device, the paper strip where a reaction mix is stored and the readout reaction takes place and finally our low-cost fluorescence detector "Lightbringer" that performs the readout measurement. Images are clickable and linked to the corresponding wiki subsection.  
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designation.
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<a href="#equation1">Equation 1</a> is motivated from the equation
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<div class="equationDiv"><img class="largeEquation" src="https://static.igem.org/mediawiki/2017/4/4b/T--Munich--Hardware_equation2.png"><span>(2)</span></div>
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<p>
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which is given in the data sheet of the LDR. The denominator is the decadic logarithm of the fraction of two light
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intensities of 100 Lx and 10 Lx. <i>R<sub>10</sub></i> and <i>R<sub>100</sub></i> are the corresponding resistances at these light intensities. The used resistor with the type designation GL5516 NT00183 has a parameter γ of 0.8.
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</p>
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<p>
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The response of a LDR depends on the wavelength λ of the incoming light. The data sheet provides information on
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the relative response normalized to the maximal response. The relative response is maximum for a wavelength of 540
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nm and is therefore appropriate for detection of green fluorophores.
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</p>
 
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<font size=7 color=#51a7f9><b style="color: #51a7f9">Hardware</b></font>
 
</td>
 
</td>
 
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<td  colspan = 6 align=center valign=center>
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<p class="introduction">
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The liberation of diagnostic tests from expensive lab infrastructure requires innovative ways of sample processing and measuring. We therefore developed a set of portable hardware tools with the goal of providing an automated sample-to-answer solution. The heart of our system is <a class="myLink" href="https://2017.igem.org/Team:Munich/Hardware/Detector">‘Lightbringer’</a>, our fluorescence detector, which is capable of measuring kinetics of biological or chemical reactions on <a class="myLink" href= "https://2017.igem.org/Team:Munich/Hardware/Paperstrip" >paper.</a> Built from 3D–printed parts and standard electronic components, it can be assembled for less than 15$, while offering a sensitivity competitive to commercial fluorescence readers. Additionally, tackling the challenge of sample pre-processing in field, we developed a portable <a class="myLink" href="https://2017.igem.org/Team:Munich/Hardware/SampleProcessing">  fluidic system</a>, featuring a temperature control unit for lysis and isothermal PCR. Conceiving a platform independent of lab infrastructure, we demonstrate the feasibility of <a class="myLink" href="https://2017.igem.org/Team:Munich/Hardware/QuakeValve"> controlling fluid flow</a> with bike tires and air balloons. All hardware components are designed and documented with the aim of enabling the community to reproduce and extend our set of tools.
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<h4>Circuit for resistance measurements</h4>
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<img width=960 src="https://static.igem.org/mediawiki/2017/2/26/Schema_final_lowres.png">
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A voltage divider as shown in the figure bellow is the simplest way to measure resistance.
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Applying Kirchhoff’s laws we get
 
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<div class="equationDiv"><img class="largeEquation" src="https://static.igem.org/mediawiki/2017/b/b1/T--Munich--Hardware_equation3.png"><span>(3)</span></div>
 
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<p>
 
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and
 
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</p>
 
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<div class="equationDiv"><img src="https://static.igem.org/mediawiki/2017/9/99/T--Munich--Hardware_equation4.png"><span>(4)</span></div>
 
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<p id="equation5">
 
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<i>R<sub>LDR</sub></i> and <i>U<sub>LDR</sub></i> are the resistance and voltage drop at the LDR. <i>R<sub>ref</sub></i> and <i>U<sub>ref</sub></i> are the resistance and voltage drop at a reference resistor. <i>U<sub>0</sub></i> is the supply voltage which we choose to be 5V. This gives
 
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</p>
 
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<div class="equationDiv"><img class="largeEquation" src="https://static.igem.org/mediawiki/2017/c/c7/T--Munich--Hardware_equation5.png"><span>(5)</span></div>
 
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<p>
 
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an equation to calculate <i>R<sub>LDR</sub></i> from <i>U<sub>LDR</sub></i>, which can be measured with the micro controller.
 
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</p>
 
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<p>
 
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We need to find an equation to choose an optimal resistor <i>R<sub>ref</sub></i> . We want a maximum change of <i>U<sub>LDR</sub></i> for a certain detection range of <i>R<sub>LDR</sub></i>. Therefore <a href="#equation5">equation 5</a> is solved for <i>U<sub>LDR</sub></i> giving
 
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</p>
 
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<div class="equationDiv"><img class="largeEquation" src="https://static.igem.org/mediawiki/2017/3/3f/T--Munich--Hardware_equation6.png"><span>(6)</span></div>
 
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<p>
 
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The change ∆<i>U<sub>LDR</sub></i> of <i>U<sub>LDR</sub></i> between a maximum value <i>R<sub>max</sub></i> and a minimum value <i>R<sub>min</sub></i> of <i>R<sub>LDR</sub></i> is
 
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</p>
 
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<div class="equationDiv"><img class="largeEquation" src="https://static.igem.org/mediawiki/2017/9/9b/T--Munich--Hardware_equation7.png"><span>(7)</span></div>
 
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<p id="equation8">
 
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which has a maximum for
 
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</p>
 
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<div class="equationDiv"><img src="https://static.igem.org/mediawiki/2017/c/c0/T--Munich--Hardware_equation8.png"><span>(8)</span></div>
 
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<p>
 
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We expect an <i>R<sub>max</sub></i> of approximately 2 MΩ and an <i>R<sub>min</sub></i> of approximately 1 MΩ. By using <a href="#equation8">equation 8</a> as a guideline we choose <i>R<sub>ref</sub></i> to be 1.5 MΩ.
 
 
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Latest revision as of 09:14, 9 December 2017

Description

Design

Demonstrate

Measurement

Applied Design

Final Results

Achievements

Protocols

Notebook

Safety

Parts

Improved Part

Part Collection

InterLab

Model

Software

Collaboration

Detector

Quake Valve

Paperstrips

Sample Processing

Silver

Gold

Engagement

Collaborations

Team members

Sponsors

Attributions

Gallery

Complete overview of all modular hardware parts in our pathogen detection system. Shown are counterclockwise and starting in in the upper left corner: The Quake valve that controls fluid flow, our sample processing device, the paper strip where a reaction mix is stored and the readout reaction takes place and finally our low-cost fluorescence detector "Lightbringer" that performs the readout measurement. Images are clickable and linked to the corresponding wiki subsection.

Hardware

The liberation of diagnostic tests from expensive lab infrastructure requires innovative ways of sample processing and measuring. We therefore developed a set of portable hardware tools with the goal of providing an automated sample-to-answer solution. The heart of our system is ‘Lightbringer’, our fluorescence detector, which is capable of measuring kinetics of biological or chemical reactions on paper. Built from 3D–printed parts and standard electronic components, it can be assembled for less than 15$, while offering a sensitivity competitive to commercial fluorescence readers. Additionally, tackling the challenge of sample pre-processing in field, we developed a portable fluidic system, featuring a temperature control unit for lysis and isothermal PCR. Conceiving a platform independent of lab infrastructure, we demonstrate the feasibility of controlling fluid flow with bike tires and air balloons. All hardware components are designed and documented with the aim of enabling the community to reproduce and extend our set of tools.