Balb/c mice were immunized intramuscularly with preS VLP (n = 6), recombinant preS protein (n = 6), or PBS (n = 6). On day 70, HBV replication was induced by hydrodynamic injection of pT-HBV1.3 plasmid via tail vein (10 μg per mouse). (A) Liver-associated HBV RNA copies were measured by qRT-PCR. (B) Immunohistochemistry analysis of liver tissues. Quantifications represent average numbers of HBcAg-positive cells per 100 × field (n = 3 with 5 fields counted per sample). Representative images are shown. Magnification, 100 ×. Serum analysis for HBsAg (C), HBeAg (D) and anti-preS antibody titers (E) was completed by ELISA on days 0, 2, 4, and 7 post transfection. (F) Serum alanine aminotransferase (ALT) activity was determined with a Hitachi 7600 Autom atic Biochemistry Analyzer. All values are presented as the average from each group, and error bars indicate ± SEM.