If somebody wants to know if they have become infected by the new coronavirus, they take a blood test which is later centrifuged and the solid and liquid parts separated obtaining a liquid known as the serum, where there are antibodies produced in response to the virus. Then the plasma is analyzed with the ELIPSE-COL test.
The ELISA type test is carried out to discover antibodies (immune system molecules) of a person that has been in contact with a pathogen, for instance, bacteria, protozoa, or viruses, such as the SARS-CoV-2.
If the person has a high concentrate of antibodies stimulated by COVID-19, the color of the sample will be intense and in case of no antibodies present, it will be colorless.
UNal Department of Chemistry Professor Luz Mary Salazar and Department of Pharmacy Professor José Manuel Lozano, both with a Ph.D. in chemistry, reported the discovery of multiple compounds, possible candidates for a diagnostic test. Some of the INS researchers reported that the specificity and sensibility of the method was 91%, that they have analyzed more than 500 samples up to now, and that one of the components reacts specifically in asymptomatic patients, but they are still working to understand why.
The researchers indicate three opportunities for this diagnostic test:
UNal researchers were after the trace of the new coronavirus genome before it arrived in Colombia, and to use their more than 30 years of experience in producing peptides capable of identifying the virus, and inclusively with the potential of becoming candidates for a vaccine.
The professors say they had the advantage that the genetic code of the virus was rapidly known. On March 18th, 12 days after the first reported case in Colombia, the INS revealed the first genetic sequence of the virus in the country. Today there are more than 1,700 reported genomes across the world.
Lozano reveals that the reported genomes are 97% identical, but that 3% difference means many mutations: “What we did was analyze 30,000 SARS-CoV-2 base pairs and looked for the antigenic determinants in 10 open reading frames (ORF), in other words, the places of the virus that the immune system could recognize as something strange and defend itself.”
While there are two lineages or “genetic variants” of the virus, in Colombia there are 12 sub-lineages, in other words, the virus that affects Colombians is not the same that in other countries, therefore it is key to have diagnostic methods engineered for “local viruses” as well as for global SARS-CoV-2 genetic variants.
Salazar says it is very difficult to synthesize and provide the immune system a complete protein, therefore we use smaller fragments known as peptides: “we discovered some that the body recognizes and produces defenses; the way we reached these molecules was by analyzing the genetic information of the virus reported around the world”.
Lozano adds that they analyzed how the cells of the immune system behave and used bioinformatic and enzymology methods to discover these molecules that the body would recognize as foreign.
After four weeks of rigorous molecular engineering, where they had to purchase reagents and mice from their pocket, they came up with the first peptide batch. They began a search to test in human serum samples and then contacted the INS, who accepted to test the peptide antigens along with some they had already developed in their lab with the ELISA technique. The goal was to create a diagnostic test with high specificity for the virus to avoid false positives and help reduce the expense of the test bought outside of the country.
Scientists Lozano, Salazar, and Organic Chemist Angela Torres tested 15 compounds in INS samples and envisioned a potential diagnostic method. With tests on mice, they discovered they produced an excellent immune response. To protect these findings, the researchers sought the help of the UNal Intellectual Property Team to submit a patent request to the United States Patent and Trademark Office (USPTO).
Read more: Clinical testing and the importance of finding a treatment for COVID-19 (in Spanish).
INS Parasitology Group Researcher Adriana Arévalo says that the ELIPSE-COL test identifies IgG (immunoglobulin G) antibodies and emphasizes that they often appear between 10 and 30 days after infection. For Dr. Lozano these tests will be key in seroprevalence studies, in other words, to become cognizant of how many people had contact with the virus and to determine how these antibodies were produced: “the serum and plasma of a person that recovered from COVID-19 will be very important to know which proteins triggered the development of antibodies”.
The researchers say that if they want to perform diagnostics from UNal, they have enough peptide antigens to assess around 2 million people, or the equivalent to 4 or 5g of molecules. They need to do a triplicate for every patient, i.e. a box with 96 well plates can have between 30 and 40 samples; this triplicate has an approximate cost of Col $200-300 (US $0.06- $0.08) per person, while the commercial tests available externally can cost up to Col $ 800 (US $0.22),” said Lozano.
The test could be scaled as there are complete areas to do so and inclusively produce 10g of the peptide, sufficient for 5 million people.
Salazar says that of the 145 candidates in the world, possible this project is the last on the list, while Lozano adds that what they have until now is that in mice it has produced an immune response. “The following step would be a controlled experiment to assess that the mice obtained antibodies and the neutralization of the virus, in other words, the virus is placed presence of human healthy cells in presence and absence of the antibodies to see if they are capable of blocking the infectious process in vitro, and that would get us closer to having a candidate for a vaccine,” although he says it is not a short-term goal.
* The Journal Vaccine No. 8 (2020) will publish the experimental data of the findings of the UNal research group.
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