We examined Epstein-Barr virus (EBV)-specific antibodies in serum samples from 64 and 59 patients with EBV-positive and -negative gastric carcinomas, respectively, and 73 healthy controls using immunofluorescence assays. EBV capsid antigen (VCA) IgG and EBV-determined nuclear antigen (EBNA) IgG were detected in all 196 subjects. The geometric mean titer (GMT) of VCA-IgG, but not EBNA-IgG, was higher in EBV-positive carcinoma cases than in EBV-negative carcinoma cases (P < 0.001). The seroprevalence rates of VCA-IgA and EBV early antigen (EA) IgG were higher in EBV-positive carcinoma cases than in EBV-negative carcinoma cases. Odds ratios (ORs) comparing seroprevalence rates between EBV-positive and -negative carcinoma cases were 3.4 (95% confidence interval [CI] = 1.3-8.8) and 6.6 (95% CI = 2.7-16.3) for VCA-IgA and EA-IgG, respectively. These results suggest that EBV reactivation occurs in vivo, since more than 90% of Japanese are infected with EBV in early childhood. The GMT of VCA-IgG in EBV-negative carcinoma cases was higher than that of healthy controls (P = 0.028). The seroprevalence rates of EA-IgG were greater in EBV-negative carcinoma cases than in healthy controls (OR = 4.9, 95% CI = 1.2-19. 7). VCA-IgA was the only antibody that showed a significantly high seroprevalence and GMT in EBV-positive carcinoma cases, but not in EBV-negative carcinoma cases. Thus, VCA-IgA can be a marker of immune response to EBV in EBV-positive carcinoma cases. Our findings support the hypothesis that if EBV is involved in the development of EBV-positive gastric carcinoma, the EBV reactivation occurs in vivo. Copyright 2000 Wiley-Liss, Inc.