DIAGNOSIS OF HEPATITIS-C VIRUS (HCV) INFECTION BY ANTIGEN-CAPTURING ELISA

Background: Hepatitis C virus (HCV) is a major cause of non-A non-B he patitis. Detection of circulating antibodies against HCV by enzyme-lin ked immunosorbent assay (ELISA) has provided the main approach for the diagnosis of HCV infection. Most ELISA kits use a mixture of core, NS 3, NS4 and NS5 antigen as capture antigens and enzyme-labeled goat ant i-human Ige as conjugate. Objectives: To establish an ELISA system bas ed on the antigen-capturing principle, using a recombinant chimeric po lyprotein containing four HCV antigenic components as antigen. Study d esign: HCV antigens were expressed in Escherichia coli as chimeric pol yprotein either in inclusion bodies or in soluble form. Protein expres sed in inclusion bodies was used as solid-phase antigen, and the antig en expressed in a soluble form was used as enzyme conjugate after bein g labeled with horseradish peroxidase (HRP). Results: Genes coding HCV antigens were cloned and sequenced, chimeric polyproteins containing four immunodominant components (core, NS3, NS4 and NS5) were expressed in E. coli both in soluble and in inclusion body form. These two chim eric proteins retained the antigenicity of HCV antigens. Antibody-capt uring ELISA using the chimeric antigens showed a sensitivity of 97% (9 7/100) and a specificity of 98% (97/99) using the reference panel from the National Institute for the Control of Pharmaceutic and Biological Products of China (NICPBC); the same assay showed a sensitivity of 97 .9% (48/49) and a specificity of 100% (43/43) using the self-establish ed reference panel. Antigen-capturing ELISA was set up using the antig en labeled with horseradish peroxidase as conjugate, and was shown to be as sensitive as (97.9%) and more specific than (100%) antibody-capt uring ELISA using the reference panel in this work. The antigen-captur ing ELISA also showed a high accordance (98.9%) with UBI HCV enzyme im munoassay (EIA) 4.0 kits (United Biomedical Inc. USA). Conclusion: Ant igen-capturing ELISA provided a convenient, sensitive and more specifi c approach for the diagnosis of hepatitis C virus infection.