Use of a novel Hepatitis C major epitope chimeric polypeptide for the diagnosis of HCV viral infection.

ABSTRACT: The genome of hepatitis C virus (HCV) consists of seven functional regions: the core, E1, E2/NS1, NS2, NS3, NS4, and NS5 regions. The U. S. Food and Drug Administration-licensed 2.0G immunoassay for the detection of anti-HCV uses proteins from the core, NS3, and NS4 regions (McHutchinson et al., Hepatology 15:19-25, 1992). The 3.0G enzyme-linked immunosorbent assay includes the protein from the NS5 region (Uyttendaele et al., Vox Sang. 66:122-129, 1994). The necessity of detecting antibodies to viral envelope proteins (E1 and E2) and to different genotype samples has been demonstrated previously (Chien et al., Lancet 342:933, 1993; Lok et al., Hepatology 18:497-502, 1993). In this study we have attempted to improve the sensitivity of the anti-HCV assay by developing a single multiple-epitope fusion antigen (MEFA; MEFA-6) which incorporates all of the major immunodominant epitopes from the seven functional regions of the HCV genome. A nucleic acid sequence consisting of proteins from the viral core, E1, E2, NS3, NS4, and NS5 regions and different subtype-specific regions of the NS4 region was constructed, cloned, and expressed in yeast. The epitopes present on this antigen can be detected by epitope-specific monoclonal and polyclonal antibodies. In a competition assay, the MEFA-6 protein competed with 83 to 96% of genotype-specific antibodies from HCV genotype-specific peptides. This recombinant antigen was subsequently used to design an anti-HCV chemiluminescent immunoassay. We designed our assay using a monoclonal anti-human immunoglobulin G antibody bound to the solid phase. Because MEFA-6 is fused with human superoxide dismutase (h-SOD), we used an anti-human superoxide dismutase, dimethyl acridinium ester-labeled monoclonal antibody for detection. Our results indicate that MEFA-6 exposes all of the major immunogenic epitopes. Its excellent sensitivity and specificity for the detection of clinical seroconversion are demonstrated by this assay.

Chien D., Arcangel P., Medina M.A., Coit D., Baumeister M., Nguyen S., George-Nascimento C., Gyenes A., Kuo G. and Valenzuela P. Use of a novel Hepatitis C major epitope chimeric polypeptide for the diagnosis of HCV viral infection. J. Clin. Microbiol., 37 (5), 1393-1397 (1999).

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