The availability of stored sera that had been obtained for other investigations permitted examination of the sequential development of hepatitis B virus (HBV) infection markers and alpha-fetoprotein (AFP) in persons who later developed primary hepatocellular carcinoma (PHC). 9 of the 11 PHC subjects had markers of HBV infection. The results showed that HBV infection could have occurred as little as 10 years before the recognition of PHC. Seroconversion from HBeAg to anti-HBe and significant rises of AFP levels occurred as long as 6 years and 2 years, respectively, before clinical onset of PHC. The finding that anti-HBe seems to precede the rise in AFP by several years suggests that in populations with a high incidence of HBV-associated PHC, or in families with high risk of PHC, the best use of AFP screening may be to monitor HBsAg carriers after seroconversion from HBeAg to anti-HBe.