Infection by hepatitis B virus(HBV) results in acute and chronic liver damages in humans. Liver products of broilers as a primary food consumed in our daily life have a close connection with public health. The prevalence of the virus in livers and serum of broilers is of great significance, owning to the potential transmission between chickens and humans. Liver tissues and serum samples were tested to investigate the prevalence of hepatitis B virus infection in slaughtered broilers, for expression of HBV antigens and antibodies. The distribution and positive rate of hepatitis B surface antigen(HBs Ag), hepatitis B core antigen(HBc Ag) and hepatitis B e antigen(HBe Ag) in liver samples were examined using immunohistochemistry. HBs Ag was mainly located in the cytoplasm of hepatocytes with a positivity of 81.61% whereas HBe Ag and HBc Ag were primarily located in the nucleus of hepatocytes with a positivity of 40.13 and 49.10%, respectively. Enzyme-linked immunosorbent assay(ELISA) analysis of serum for HBV serological markers demonstrated a high prevalence of hepatiits B surface antibody(HBsA b, 54.91%) and hepatitis B core antibody(HBcA b, 27.68%), whereas HBeA b, HBsA g and HBeA g were rarely detectable. Classic hepatitis pathological changes, including swollen hepatocytes, focal parenchymal necrosis, lymphocytic infiltration and hyperplasia of fibrous connective tissues were observed using histopathological analysis. Some of the liver samples were found positive for HBV DNA using nested PCR. Sequence comparison confirmed that all sequences shared 97.5–99.3% identity with human HBV strains. These results demonstrated the existence of HBV in livers and serums of broilers. Animals or animal products contaminated with HBV could raise an important public health concern over food safety and zoonotic risk.