Identification of a domain containing B-cell epitopes in hepatitis C virusE2 glycoprotein by using mouse monoclonal antibodies

Evidence from clinical and experimental studies of human and chimpanzees su ggests that hepatitis C virus (HCV) envelope glycoprotein E2 is a key antig en for developing a vaccine against HCV infection. To identify B-cell epito pes in HCV E2, six murine monoclonal antibodies (MAbs), CET-1 to -6, specif ic for HCV E2 protein were generated by using recombinant proteins containi ng E2t (a C-terminally truncated domain of HCV E2 [amino acids 386 to 693] fused to human growth hormone and glycoprotein D). We tested whether HCV-in fected sera were able to inhibit the binding of CET MAbs to the former fusi on protein. Inhibitory activity was observed in most sera tested, which ind icated that CET-1 to -6 were similar to anti-E2 antibodies in human sera wi th respect to the epitope specificity. The spacial relationship of epitopes on E2 recognized by CET MAbs was determined by surface plasmon resonance a nalysis and competitive enzyme-linked immunosorbent assay. The data indicat ed that three overlapping epitopes were recognized by CET-1 to -6. For mapp ing the epitopes recognized by CET MAbs, we analyzed the reactivities of CE T MAbs to six truncated forms and two chimeric forms of recombinant E2 prot eins. The data suggest that the epitopes recognized by CET-1 to -6 are loca ted in a small domain of E2 spanning amino acid residues 528 to 546.