Hypervariable region 1 (HVR1), located in the N-terminal region of a putative second envelope glycoprotein (gp70) of hepatitis C virus (HCV), contains immunological B-cell epitopes which might be neutralizing epitopes. To clarify whether B-cell epitopes within HVR1 are common among virus isolates or specific for the homologous virus isolate, we examined the reactivities of sera from 53 patients with chronic hepatitis or hepatocellular carcinoma/liver cirrhosis against two different HVR1 peptides (HVR1 I-1 and HVR1 Y-1) derived from patient I with sporadic acute hepatitis and an asymptomatic carrier Y, respectively, using our original assay system for the detection of anti-HVR1 antibody. All patients examined had a history of blood transfusion. Most sera showed no reactivity with either HVR1 I-1 or HVR1 Y-1 peptide. Only seven and fourteen serum samples reacted significantly, although weakly, with HVR1 I-1 and HVR1 Y-1 peptides, respectively, compared with the serum from patient I or asymptomatic carrier Y. The blood transfusions of most reactive cases had occurred more than thirty years earlier. Six cases reacted with both HVR1 I-1 and HVR1 Y-1 peptides, but further analysis revealed that only three cases reacted weakly with the peptide for either epitope I or II, identified within HVR1 I-1. These results indicate that the B-cell epitopes within HVR1 are fairly specific for the homologous virus isolate, and this may represent a serious difficulty in the development of a vaccine against HCV.