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Team:Linkoping Sweden/Project/Biology
From 2014.igem.org
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<p>Our approach is to use an antibody or functionally similar structure in combination with fluorescence probes to detect the Ara h antigen. This system will have three potential parts: an antibody complex consisting of an polyclonal antibody connected to a fluorescent probe, a default-epitope complex consisting of five epitopes linked to a fluorescent probe, and the Ara h protein. These are potential parts in the sense that system will always contain the first two parts (antibody complex and the default epitope complex), but will only contain the third part, Ara h protein, if the inserted sample has been contaminated with peanut protein. The biosensor system is to be implemented in a device consisting of a box containing the sensors necessary to detect fluorescence and any eventual changes in fluorescence. This information is then processed by a microprocessor to give a color indication, red or green, correlating to the changes in fluorescence. So how exactly will the system work? Let’s break it down step-by-step:</p> | <p>Our approach is to use an antibody or functionally similar structure in combination with fluorescence probes to detect the Ara h antigen. This system will have three potential parts: an antibody complex consisting of an polyclonal antibody connected to a fluorescent probe, a default-epitope complex consisting of five epitopes linked to a fluorescent probe, and the Ara h protein. These are potential parts in the sense that system will always contain the first two parts (antibody complex and the default epitope complex), but will only contain the third part, Ara h protein, if the inserted sample has been contaminated with peanut protein. The biosensor system is to be implemented in a device consisting of a box containing the sensors necessary to detect fluorescence and any eventual changes in fluorescence. This information is then processed by a microprocessor to give a color indication, red or green, correlating to the changes in fluorescence. So how exactly will the system work? Let’s break it down step-by-step:</p> | ||
<p>There are two possible scenarios for our biosensor system, either the sample does not contain peanut protein (Ara h) (Fig. 1A) or it does (Fig. 1B). The first scenario is default for our biosensor; it contains two interacting parts in the form of an antibody connected to a fluorescent probe (Fluorescein isothiocyanate, or FITC) and five epitopes bound to a fluorescent probe (Red Fluorescence Protein or RFP). The epitopes binds to the antibody, and when a light source is shone on the biosensor the incoming light is absorbed by FITC and transferred to RFP through Förster Resonance Energy Transfer (FRET) (Fig. 2). The emitted wavelength is detected and then processed by a microprocessor which makes a LED light-panel shine green. Green light thus indicates that the system only contains the antibody and epitope complexes, and has not been contaminated by peanut protein.</p> | <p>There are two possible scenarios for our biosensor system, either the sample does not contain peanut protein (Ara h) (Fig. 1A) or it does (Fig. 1B). The first scenario is default for our biosensor; it contains two interacting parts in the form of an antibody connected to a fluorescent probe (Fluorescein isothiocyanate, or FITC) and five epitopes bound to a fluorescent probe (Red Fluorescence Protein or RFP). The epitopes binds to the antibody, and when a light source is shone on the biosensor the incoming light is absorbed by FITC and transferred to RFP through Förster Resonance Energy Transfer (FRET) (Fig. 2). The emitted wavelength is detected and then processed by a microprocessor which makes a LED light-panel shine green. Green light thus indicates that the system only contains the antibody and epitope complexes, and has not been contaminated by peanut protein.</p> | ||
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<div class="image-box" style="width:400px"> | <div class="image-box" style="width:400px"> | ||
<a href="https://static.igem.org/mediawiki/2014/0/09/Linkoping_sweden_GFP_RFP_FRET.png" target="blank"> | <a href="https://static.igem.org/mediawiki/2014/0/09/Linkoping_sweden_GFP_RFP_FRET.png" target="blank"> | ||
<img src="https://static.igem.org/mediawiki/2014/0/09/Linkoping_sweden_GFP_RFP_FRET.png" width="400px" height="auto" title="FRET"> | <img src="https://static.igem.org/mediawiki/2014/0/09/Linkoping_sweden_GFP_RFP_FRET.png" width="400px" height="auto" title="FRET"> | ||
</a> | </a> | ||
| - | <p>FRET (Click to enlarge).</p> | + | <p>Fig 1. FRET (Click to enlarge).</p> |
</div> | </div> | ||
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<p>The other possible scenario is that the inserted food sample has been contaminated by peanut and that system thus contains Ara h. The peanut protein will bind with a higher affinity to the FITC-antibody complex, thus forcing the epitope-RFP complex out of the picture. Ara h bound to the antibody-FITC complex reduces FRET as applied light is absorbed and emitted solely by FITC. This results in a wavelength change which is detected and processed, causing in a LED light-panel change from green to red. Red light thus indicates that the system has been contaminated by peanut and that the food is unsafe to consume. | <p>The other possible scenario is that the inserted food sample has been contaminated by peanut and that system thus contains Ara h. The peanut protein will bind with a higher affinity to the FITC-antibody complex, thus forcing the epitope-RFP complex out of the picture. Ara h bound to the antibody-FITC complex reduces FRET as applied light is absorbed and emitted solely by FITC. This results in a wavelength change which is detected and processed, causing in a LED light-panel change from green to red. Red light thus indicates that the system has been contaminated by peanut and that the food is unsafe to consume. | ||
</p> | </p> | ||
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<img src="https://static.igem.org/mediawiki/2014/e/ef/Linkoping_sweden_mechanism.png" width="400px" height="auto" title="Mechanism"> | <img src="https://static.igem.org/mediawiki/2014/e/ef/Linkoping_sweden_mechanism.png" width="400px" height="auto" title="Mechanism"> | ||
</a> | </a> | ||
| - | <p>Mechanism (Click to enlarge).</p> | + | <p>Fig 2. Mechanism (Click to enlarge).</p> |
</div> | </div> | ||
</div> | </div> | ||
Revision as of 08:03, 14 October 2014
The Biology
Our approach is to use an antibody or functionally similar structure in combination with fluorescence probes to detect the Ara h antigen. This system will have three potential parts: an antibody complex consisting of an polyclonal antibody connected to a fluorescent probe, a default-epitope complex consisting of five epitopes linked to a fluorescent probe, and the Ara h protein. These are potential parts in the sense that system will always contain the first two parts (antibody complex and the default epitope complex), but will only contain the third part, Ara h protein, if the inserted sample has been contaminated with peanut protein. The biosensor system is to be implemented in a device consisting of a box containing the sensors necessary to detect fluorescence and any eventual changes in fluorescence. This information is then processed by a microprocessor to give a color indication, red or green, correlating to the changes in fluorescence. So how exactly will the system work? Let’s break it down step-by-step:
There are two possible scenarios for our biosensor system, either the sample does not contain peanut protein (Ara h) (Fig. 1A) or it does (Fig. 1B). The first scenario is default for our biosensor; it contains two interacting parts in the form of an antibody connected to a fluorescent probe (Fluorescein isothiocyanate, or FITC) and five epitopes bound to a fluorescent probe (Red Fluorescence Protein or RFP). The epitopes binds to the antibody, and when a light source is shone on the biosensor the incoming light is absorbed by FITC and transferred to RFP through Förster Resonance Energy Transfer (FRET) (Fig. 2). The emitted wavelength is detected and then processed by a microprocessor which makes a LED light-panel shine green. Green light thus indicates that the system only contains the antibody and epitope complexes, and has not been contaminated by peanut protein.
Fig 1. FRET (Click to enlarge).
The other possible scenario is that the inserted food sample has been contaminated by peanut and that system thus contains Ara h. The peanut protein will bind with a higher affinity to the FITC-antibody complex, thus forcing the epitope-RFP complex out of the picture. Ara h bound to the antibody-FITC complex reduces FRET as applied light is absorbed and emitted solely by FITC. This results in a wavelength change which is detected and processed, causing in a LED light-panel change from green to red. Red light thus indicates that the system has been contaminated by peanut and that the food is unsafe to consume.
Fig 2. Mechanism (Click to enlarge).
