Difference between revisions of "Team:INSA-Lyon/Description"

Technological choices

Biomarkers detection is a field largely explored both by iGEM teams and industrials. Mostly used techniques rely on antibodies. Developing antibodies-based tests has some drawbacks: lengthy and pricey process of identification and production, short shelf-life… innovations are needed to develop more frugal tests.

Our main innovation relies on the use of aptamers technology. Aptamers are short single stranded nucleic acids. They basically act as antibodies. They have numerous advantages. Compared to antibodies, the processes of selection and production of aptamers are quicker, easier, and cheaper. Aptamers can be synthesized by pure chemistry, whereas antibodies need complex eukaryotic chassis to be produced. Aptamers are an user-friendly technology! They also have the advantage to be easily chemically modifiable. Three kinds of BioBricks were constructed and sent to the registry to build our detection device:
target generators (protein subunits of the HIV-1 Reverse Transcriptase and HBsAg, biomarker of Hepatitis B, deposited parts BBa_K1934060 and BBa_K1934061)
anchor allowing the fixation of the aptamer to the paper: our biological detection system needs to be anchored on paper. An existing fusion protein (part BBa_K1499004) could do that, but it was not characterised, so we improved it and tested it fully.

Learn more about aptamers

How did we do it?

Two approaches were developed during this project. First, a detection system based on fluorescence. It worked well, but we realized it could not be used at home. The second approach was a detection system based on latex beads.

A first aptamer is covalently linked to the latex beads. After adding the sample onto the beads, they migrate on a paper strip. On this paper strip there are also complementary aptamers which recognize a different epitope of the biomarker. If the target is present in the sample, it will be taken in sandwich between the aptamers on the beads and the fixed aptamers. A visible black band will progressively appear, due to the beads’ coloration. If the target is not present, beads will be unable to form this sandwich and won't be stopped.

A control is also needed. So after the detection band there is also a control band made with complementary strands of the DNA on the beads. Just by base-pairing the beads will be stopped. They are put in excess for the detection, so even if there is the target in the sample to be analyzed, some beads are going to migrate to the control band.

How to fix aptamers on the paper strip? We got inspired by another iGEM project, from the 2014 Stanford-Brown-Spelman team. They developed a Streptavidin linked to a Cellulose binding domain, fusion protein. We took this idea and pushed it a little further, characterizing their part and building new and more efficient ones. Basically, aptamers are biotinylated, they bind to the Streptavidin part, which is fixed onto cellulose thanks to the cellulose binding domains.

Another important aspect of the project was the design of a real device. If one wants to sell a detection platform, it must not only be the biological part, but also a real device to pack it. So we designed and 3D printed a device. A first model was designed as a prototype to test our paper strips. A second one was designed to be specifically used in real life. It’s a cornerstone for our project, because that’s what people are going to see and use.

Sources of Inspiration and motivations

Aptamers were not used at all in the labs of our school, but we learned they existed by reading literature. We immediately found out it was a promising technology. We looked further in literature to get some ideas about signal transduction with aptamers. That brought us to the idea of using fluorescence. After some tests, we were a little disappointed, so we went to an international congress about Aptamers in Bordeaux (France). Some researchers, and particularly Dr. Ciara O’Sullivan, brought us to the idea of using nanoparticles covalently linked to the aptamers. This idea seduced us and we immediately started working on it. But instead of using gold nanoparticles like most of the labs we chose to use latex beads. That’s a much more eco-friendly option, plus on a large scale it’s less expensive.

Our major motivation was to provide the world with a good solution to detect STIs. But we soon realized that what we achieved is actually a much more general strategy for developing quick and efficient tests. Indeed we developed a strategy which is extendable to pretty much every target you can think about. So we could imagine to sell the device and paper strips to be put inside the casing as a platform to be adapted with aptamers for each situations. Instead of detecting STIs we could detect water pollutants (heavy metals, pesticides…), other health issues (cancer, bacterial infections...) and food contamination. So we hope some future iGEM teams will follow us on this exciting path!