Team:WPI-Worcester/Proof-of-Principle

From 2014.igem.org

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<p><h9>Microscopy</h9></p><p>Content goes here</p><p><center><img src="microscopy images"/><img src="microscopy images"/></center></p>
<p><h9>Microscopy</h9></p><p>Content goes here</p><p><center><img src="microscopy images"/><img src="microscopy images"/></center></p>
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<p><h9>Agglutination Assay</h9></p><p>We were able to successfully achieve the results we wanted with the Agglutination Assay.  When antibodies are in the presence of their matching antigen pairs, they will bind to each other.  Antibodies have two antigen binding sites which allows a network of antibody-antigen expressing bacteria to form.  This network manifests on a large scale as a mat of bacteria on the bottom of the containing vessel, as opposed to a solid dot formed by unagglutinated antibodies. We based our assay off of <a href="http://www.plosntds.org/article/info%3Adoi%2F10.1371%2Fjournal.pntd.0001946">this paper</a>. <br>
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<p><h9>Agglutination Assay</h9></p><p>We were able to successfully achieve the results we wanted with the Agglutination Assay.  When antibodies are in the presence of their matching antigen pairs, they will bind to each other.  Antibodies have two antigen binding sites which allows a network of antibody-antigen expressing bacteria to form.  This network manifests on a large scale as a mat of bacteria on the bottom of the containing vessel, as opposed to a solid dot formed by unagglutinated antibodies. We based our assay off of <a href="http://www.plosntds.org/article/info%3Adoi%2F10.1371%2Fjournal.pntd.0001946">this paper</a>.</p>
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For our proof of principle agglutination, we used surface-bound YFP(Yellow Fluourescent Protein) and a corresponding GFP(Green Fluourescent Protein)/YFP antibody (the part of the protein that the antibody binds to is not affected by the part that causes the color change.) We used both of the surface expression proteins we had at our disposal: BclA (the one we made) and INP (a pre-existing biobrick). As controls, we used internally expressed YFP as well as a protein that was extrnally expressed but did not match with the antibody (we used the CAEV protein we made.) <br>
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<p>For our proof of principle agglutination, we used surface-bound YFP(Yellow Fluourescent Protein) and a corresponding GFP(Green Fluourescent Protein)/YFP antibody (the part of the protein that the antibody binds to is not affected by the part that causes the color change.) We used both of the surface expression proteins we had at our disposal: BclA (the one we made) and INP (a pre-existing biobrick). As controls, we used internally expressed YFP as well as a protein that was extrnally expressed but did not match with the antibody (we used the CAEV protein we made).</p>
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This photo clearly shows that the wells containing the bacteria with the matching antigen on the surface formed agglutinated mats while the control wells did not agglutinate and formed dots.
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<p><center><img src="https://2014.igem.org/File:WPI_AgglutinationResults.png"/></center></p>
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</p><p><center><img src="agglutination"/><img src="agglutination"/></center></p>
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<p>This photo clearly shows that the wells containing the bacteria with the matching antigen on the surface formed agglutinated mats while the control wells did not agglutinate and formed dots.
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Revision as of 04:51, 17 October 2014

Team:WPI-Worcester - 2014.igem.org

 

Team:WPI-Worcester

From 2014.igem.org


Proof of Principle

Microscopy

Content goes here

Agglutination Assay

We were able to successfully achieve the results we wanted with the Agglutination Assay. When antibodies are in the presence of their matching antigen pairs, they will bind to each other. Antibodies have two antigen binding sites which allows a network of antibody-antigen expressing bacteria to form. This network manifests on a large scale as a mat of bacteria on the bottom of the containing vessel, as opposed to a solid dot formed by unagglutinated antibodies. We based our assay off of this paper.

For our proof of principle agglutination, we used surface-bound YFP(Yellow Fluourescent Protein) and a corresponding GFP(Green Fluourescent Protein)/YFP antibody (the part of the protein that the antibody binds to is not affected by the part that causes the color change.) We used both of the surface expression proteins we had at our disposal: BclA (the one we made) and INP (a pre-existing biobrick). As controls, we used internally expressed YFP as well as a protein that was extrnally expressed but did not match with the antibody (we used the CAEV protein we made).

This photo clearly shows that the wells containing the bacteria with the matching antigen on the surface formed agglutinated mats while the control wells did not agglutinate and formed dots.