The value of two new hepatitis C virus (HCV) antigens for detection of HCV antibodies was studied. These two recombinant antigens were derived from the nonstructural-5 (NS5) and envelope-2 (E2) region of the HCV genome. In a panel of 33 HCV-RNA positive samples with indeterminate Riba-2 confirmatory test results, 29 samples (88%) showed additional antibody reactivity against E2 and 12 samples (36%) showed additional reactivity against NS5. Among 39 HCV-RNA negative, Riba-2 indeterminate donor samples, no additional E2 or NS5 reactivity was found in 34 samples (87%); while 5 samples (13%) showed additional reactivity against NS5 and/or E2. E2 reactivity thus resolved the majority of hitherto indeterminate samples. In serial samples from nine posttransfusion hepatitis C patients, NS5 and E2 antibodies did not appear earlier than classical HCV antibodies. However, E2 antibodies eventually appeared in all nine patients. The recombinant E2 might be a candidate antigen for future HCV antibody assays.