Imunodiagnóstico com Beads Single e Multiplex para detecção de anticorpos contra as proteínas recombinantes Core, Ns3, Ns4B E Ns5A do vírus da hepatite C

Abstract

Introduction: Liver infections caused by viruses are among the most serious, with prevalence worldwide. The diagnosis of HCV is made by serological tests for screening and confirmatory molecular tests. Although the diagnostic tests currently used have high levels of sensitivity and specificity, they still have false-negative and positive results, in addition to having a long workday. Objective: To develop a single and multiplex method for the detection of antibodies against the recombinant proteins CORE, Ns3, Ns4b and Ns5a of the hepatitis C virus, coupled to functionalized microbeads. Material and Methods: HCV CORE recombinant proteins, Ns3, Ns4b and Ns5a were expressed in E. coli BL21 (DE3) pLysS and purified by Ni-NTA. For single and multiplex tests, the proteins were coupled to carboxylated magnetic beads. Thirty samples of HCV + sera and 30 samples of control sera were used to analyze the recognition of antibodies against proteins coupled to beads, using the flow cytometry method. Results and conclusion: The recombinant proteins CORE, Ns3, Ns4b, Ns5a N-terminal and Ns5a C-terminal were successfully expressed in E. coli host and recovered in the soluble portion of the bacterial extract using a nickel column. The recombinant proteins coupled in magnetic beads were immunoreactive for the sera of patients with chronic hepatitis C, with high sensitivity and specificity. The multiplex assay improved the sensitivity and specificity of the tests compared to the individual assays.