Sustainable viral suppression with hepatitis B surface antigen (HBsAg) loss is the new treatment goal for chronic hepatitis B (CHB). The role of antibodies in therapies for persistent hepatitis B virus (HBV) infection has received constant attention. While immunotherapy holds great promise, challenges for the antibody-based prevention and control of HBV in CHB include broad HBV antigenic diversity and the need for long-term viral suppression. In this study, we identified a new anti-HBsAg nanobody (Nb), 125s, isolated from HBsAg-immunized alpaca and confirmed its excellent potency in HBsAg clearance and broad-spectrum therapeutic activity against three HBV subtypes in vivo. In addition, we characterized a novel epitope at the C-terminus of the HBsAg surface motif from amino acids 157 to 174. A 125s-based long-term passive immunization program was efficacious at HBsAg clearance and inducing cellular immune responses, offering a promising outlook for CHB immunotherapy.