Production of dengue virus recombinant proteins for use in rapid tests diagnostics

• Grant number: 17/22737-6
• Support Opportunities: Research Grants - Innovative Research in Small Business - PIPE
• Start date: July 01, 2018
• End date: May 31, 2019
• Field of knowledge: Biological Sciences - Biochemistry - Molecular Biology
• Principal Investigator: Maria Angélica de Camargo
• Grantee: Maria Angélica de Camargo
• Company: Wama Produtos para Laboratório Ltda
• CNAE: Fabricação de instrumentos e materiais para uso médico e odontológico e de artigos ópticos
• City: São Carlos
• Associated researchers: Gabriel Belem de Andrade ; Jessica Caldeira de Lima ; Juliana Pereira Schnetzler

Abstract

Dengue virus (DENV) is transmitted by mosquitoes of the genus Aedes and was first isolated in 1943. Almost 400 million people are infected every year with the virus, and half of the world population is subject to the geographical distribution of the virus infection. Until now, five virus serotypes (DENV-1, DENV-2, DENV-3, DENV-4 and DENV-5) were described, all of which are found in Brazil except DENV-5. Because of heterogeneous immunity, secondary infections by different serotypes are possible and relatively common. Although they may be asymptomatic, DENV infections also cause Dengue Fever (DF) and Dengue Hemorrhagic Fever/Dengue Shock Syndrome (FHD/SCD), the latter being a serious disease that can lead to death if it is not quickly diagnosed and treated. Rapid Diagnostic Tests (TRDs) are performed in a single step and are based on the interaction between antigens or antibodies present in the sample and reagents impregnated on the rapid test platform. Such tests provide a fast and efficient results to the end customer without requiring major investments in training or laboratory setting. Aiming to expand the technical expertise and the company's portfolio of products, this project aims to make feasible the development of reagents that are currently acquired by import, specifically dengue virus antigens. The viral antigens will be produced via the bacterial expression system (Escherichia coli), using nucleotide sequences of dengue virus proteins (DENV), which will be analyzed, optimized and synthesized chemically. The synthesized sequences will be cloned into the expression vector, resulting in proteins fused to a his-tag, which will be purified by affinity chromatography. The recombinant proteins obtained will be conjugated with colloidal gold and used to capture anti-dengue antibodies, composing the Rapid Diagnostic Tests (TRD) based on immunochromatography, a broad domain approach of the proposing company. This project will enable a biotechnological innovation at Wama Diagnóstica, ensuring control of research and development stages, reducing material costs and eliminating dependence on international suppliers. (AU)