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| − | <font size=7 color=#51a7f9><b style="color: #51a7f9">Hardware</b></font> | + | <font size=7 color=#51a7f9><b style="color: #51a7f9">Sample Processing</b></font> |
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| − | <p class="introduction">
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| − | The hardware device is one of the main pillars of our project. We mainly developed two separate devices, a fluorescence detector "Lightbringer" and a lysis and isothermal PCR device "Heatbringer".
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| − | </p>
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| − | </td>
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| − | <h3>Heatbringer</h3>
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| − | "Heatbringer" was developed by our Hardware crew at TECHFEST Munich. We tried to think of the easiest way of designing a heating device without the need for expensive materials. In order to achieve that, we used Peltier elements for heating the sample since this would allow us to still use rather inexpensive and simple materials such as 3D printed PLA to produce the case of the system. This is due to the characteristics of a Peltier element to create a temperature gradient.
| + | We envision CascAID as a complete sample-to-answer solution for pathogen detection. Apart from our fluorescence detector “Lightbringer”, we hence had to realize a user friendly environment for the extraction and amplification of target RNA from a patient sample via lysis, followed by a combined reverse transcription, isothermal PCR and transcription step. |
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| − | <pre style="text-align: left;">
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| − | A Peltier element uses the Peltier effect to create a temperature gradient.
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| − | The Peltier effect describes a phenomenon in which electrical current leads to a transport of heat;
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| − | therefore it is also sometimes called Heat Pump. It arises at the junction of two different conductors.
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| − | Therefore, typical Peltier elements are alternating layers of two conductors through which current is driven.
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| − | Through this, one can exploit the Peltier effect at every junction of the conductors,
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| − | creating a temperature gradient considerably higher than when only having one junction.
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| − | </pre>
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| − | <p>
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| − | The surface in contact with the case would, even at temperatures as high as 95 °C for cell lysis, not be high enough to melt the plastic since it would show a significantly lower temperature. How low this temperature gets is determined by the characteristics of the Peltier element in use.
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| − | <br>
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| − | <br>
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| − | For controlling the heating process, we regulated the on/off time of the Peltier element with a Raspberry Pi 3 using temperature sensors and a relay. We connected the Peltier elements to the relay and wrote a script that would turn on the Peltier element until the sensor has measured a certain temperature and turn it off up until the temperature drops to a given value. As a power source, we used a common power bank to make the device portable. This way, we were able to create simple and low-cost constant temperature chambers.
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| − | <br>
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| − | <br>
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| − | Into the developed isothermal chambers, we build in a PDMS chip in which we could introduce the sample and let it traverse the lysis at ca. 80 °C and the isothermal amplification at 37 °C by pumping it through the channels in the PDMS chip.
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| − | <br>
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| − | <br>
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| − | The temperature sensors are run via an I2C Bus at Bus 40 and 41 to allow connection of several sensors via one bredboard.
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| − | <br>
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| − | Fluid flow in the PDMS chip is ensured by a 3D printed peristaltic pump driven by 2 stepper motors. The circuit is shown in Figure 1. The pump was taken from Thingiverse.com (https://www.thingiverse.com/thing:254956) and adapted to our needs.
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| − | <img id="TopPicture" width="960" src="https://static.igem.org/mediawiki/2017/a/aa/T--Munich--HardwarePagePictures_Peristaltic_Pump.svg">
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| − | </p>
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| − | <p>
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| − | <i> Figure 1: Circuit diagram for a peristaltic pump using two stepper motors for pumping. </i>
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| − | </p>
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| − | <img id="TopPicture" width="960" src="https://static.igem.org/mediawiki/2017/8/85/T--Munich--HardwarePagePictures-Heatbringer.png">
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| − | <p>
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| − | <i> Figure 2: Circuit diagram for the Heatbringer Device</i>
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| | </p> | | </p> |
| | </td> | | </td> |
