Desenvolvimento de um ensaio de fluxo lateral à base de proteína p24 recombinante para diagnóstico de leucose bovina

Abstract

Enzootic Bovine Leukosis is a viral disease that causes significant loss in the production chain, in addition to being a potential risk to human health. Nevertheless, diagnostic tests available on the market are costly and time-consuming, influencing the producer's decision to diagnose the entire herd. Therefore, there is no traceability of the disease and much less an incentive to eradicate it. In view of this, we propose the development of a lateral flow assay (rapid diagnosis) based on the recombinant p24 protein as an antigen. The p24 protein is present in the nucleocapsid of the vírus and can be produced through recombinant DNA technology, in Escherichia coli. For this purpose, blood samples from infected cows were donated by local veterinarians and submitted to DNA extraction and gene amplification by PCR. Recombinant DNA based on the pUC19 vector will be transformed into E. coli. Protein purification will be performed by selective permeation, three-phase extraction and hydrophobic chromatography. Purified samples will be analyzed by HPLC-UV. The prototype test will consist of cellulose and nylon filters, and a nitrocellulose membrane in which the antigen will be conjugated with colloidal gold to visualize the lines. This project will be conducted in a partnership with the company Xavier & Loureiro Biotec LTDA, with the trade name Gene de Yebá.