Studies of HCV pathogenesis and antiviral research have been hampered by the lack of adequate cell-culture and small-animal models. The culturing of human primary hepatocytes would greatly facilitate the model development in HCV research. The availability of robust infectious virus, JFH1 (i.e., genotype 2) strain, will further increase the interest in using primary hepatocyte cultures. This cell model system will significantly enhance research in the areas of antiviral research and host-virus interaction, but obtaining pure and viable human primary hepatocytes is not trivial. We have optimized a method of liver perfusion and primary hepatocyte isolation that allows us to establish robust and reliable human primary hepatocyte cultures. Moreover, we have demonstrated that these primary cultures are susceptible to authentic HCV infection in vitro.