Hepatitis B virus (HBV) transgenic mice that are immunologically tolerant to HBV-encoded Ags represent a model of chronic HBV infection suitable for the development of therapeutic immunization strategies before testing in humans. Five lineages of HBV transgenic mice were immunized with plasmid DNA that encodes hepatitis B surface Ag (HBsAg) or with cytokine-activated bone marrow-derived dendritic cells (DCs) in an attempt to break tolerance to HBsAg at the B and T cell levels. DNA immunization stimulated an Ab response but not a cytotoxic T lymphocyte response to HBsAg in two of the five transgenic lineages studied. In contrast, infusion of activated transgenic or nontransgenic DCs stimulated a splenic CTL response in all three transgenic lineages immunized in this manner at precursor frequencies comparable to those in nontransgenic mice, indicating that DC function is normal, and HBsAg-specific CTLs are present but functionally silent in these transgenic animals. Importantly, none of the animals developed hepatitis or displayed suppressed viral gene expression or replication following either DNA immunization or DC administration even in the presence of anti-hepatitis B surface (HBs) Abs and HBs-specific CTLs. These results indicate that Ag presentation by cytokine-activated DCs can break tolerance and trigger an anti-viral CTL response in HBV transgenic mice, and they suggest that this strategy is more efficient than DNA immunization in this setting. Nonetheless, more efficient immunization strategies are needed to stimulate an immune response of sufficient quality and magnitude to achieve an immunotherapeutic antiviral effect.