Lymphocytes isolated from recipients of hepatitis B vaccine were studied for their immune response to HBsAg in vitro. Peripheral blood mononuclear cells (PBMs) from 70 to 80% of 40 vaccinees yielded proliferative indices larger than 2 after 5 to 7 days incubation with HBsAg. This in vitro proliferative response could be augmented by incubating the cells with HBsAg and supernatants of activated T cells for 2 weeks or longer. After 7 to 10 days, in vitro stimulation with antigen, PBMs (1 ± 106) could yield 5 to 15 HBsAg-specific antibody-secreting plaque-forming cells. The antibody to HBsAg produced in vitro was greatly increased in cultures that contained antigen-specific B cells enriched by panning with HBsAg-coated plates and a T cell growth factor-dependent, HBsAg-specific autologous T cell line. The results indicate that HBsAg-specific B and T cells are present, although at low frequencies, in the circulation of hepatitis B vaccinees.