Annual mass examination was performed between 1972 and 1997 in Tomie-town, Goto Islands, Japan, where hepatitis B virus (HBV) infection is very prevalent. In the present study, the incidence of spontaneous loss of hepatitis B surface antigen (HBsAg) in HBsAg carriers was determined in this area. Three thousand and nineteen inhabitants were tested for HBsAg two or more times in our annual surveys. Among them, 131 (4.3%) were defined as chronic HBsAg carriers based on the persistence of HBsAg for 1 or more years. These 131 subjects were followed for 12.2 +/- 7.6 years. During the follow-up period, spontaneous loss of HBsAg occurred in 38 (29%) of the 131 carriers, with a yearly incidence of 2.5%. This loss was seen more frequently in carriers aged 40 years or more on enrollment than in those aged less than 40 years during the same observation periods (P = 0.0141), irrespective of sex or the results of liver function tests. The values for liver function test results were similar before and after loss of HBsAg in these carriers. Stored serum samples were available for later analysis of HBV-DNA by polymerase chain reaction in 32 carriers with loss of HBsAg. The HBV-DNA sequence was detected in 26 (81%) and 2 of the 32 carriers (6%) before and after loss of HBsAg, respectively. These results indicate that spontaneous loss of HBsAg, largely attributable to clearance of viremia, occurs age-dependently in chronic carriers.