Generation of RDTs (Rapid Diagnostic Tests) for local and regional viral diseases

SANDRA GOÑI; MATÍAS LORCH; MARÍA SOLEDAD COLLADO

Cadiz

Congreso; XIV Congreso Nacional de Virologia; 2017

Sociedad Española de Virología

Objetives: The aim of laboratory is establish a workflow that converges in rapid diagnostic tests (RDTs) for serological identification of several viral infectiuos agents that have local and regional importance, with implications in diagnostic and epidemiological surveillance.Methodology: In first place is very important to recognize the important antigens for the virus of interest. In this way, are developed analysis for determine the more important epitopes (immunobioinformatics), and proceed to work with different versiones of recombinant proteins. We work with NS1 for DENV, YFV, SLEV, WNV and ZIKV Flavivirus; NS5 for SLEV Flavivirus; VP1 for human parvovirus B19; and L1 for HPV. We are use principally the bacterial plataform to produce the recombinats proteins, but we have an antigen presentation system based on Z protein of JUNV for mammals cells that can be used commplementarily. To the RTDs is necesary to purified the IgGs or MAbs (monolconal antibodies). For this, we are working in an affinity chromatography that replace the comercial adquisition, making lower the costs production.Results: Until this moment was posible to obtain the recombinant proteins in bacterial systems, fused to an histidine tag (HT), with good expression levels. The recombinant generated proteins are: HT-NS1SLEV, HT-NS1DENV-1, HT-NS1WNV, HT-NS1ZIKV, HT-NS1YFV, HT-NS1-EpSLEV, HT-NS5SLEV, HT-VP1B19, HT-L1HPV. We are work in the expression optimization and purification for several of them. Alternatively, we fused the entire sequence of NS1SLEV to a system based on the Z protein of Junín virus, obtaining VLPs that are being analyzed in their composition. Therefore, the HT-NS1SLEV protein was used to inoculate mice and evaluate the humoral response α-NS1SLEV, and start the protocol for monoclonal antibody generation. As to the affinity chromatography, the first steps for the production of affinity protein are done and then will be couple to the matrix.Conclusions: The local RDTs kits development is an important work area at cover in our countries. The availibilty of this tools at health institutions gives place to access at a correct diagnosis and to the disponibility of epidemiological data useful for planning plague control strategies, between others. In this way, we have several recombinants proteins recently produced, and we start to make a MAbs, a fundamental component that ensure the specificity and quality of the kits.