Figure 6. The Human Hepatocyte Culture System Produces Infectious HCV.

Day-5 primary human hepatocytes (1.8 million cells/60 cm²) were infected with HCV genotype 1 (56,000 HCV virions); genotype 2 (68,000 HCV virions); or genotype 3 (22,400 HCV virions) for 72 hr as described in Materials and methods. Upper panel. HCV E-2 was detected by immunopurification and immunoblotting from cell layers of naïve primary human hepatocytes infected with HCV genotypes 1, 2 or 3 for 72-hr (lanes 5–7) produced by human hepatocytes infected with HCV genotypes 1, 2 or 3 for 72-hr (lanes 2–4) as described in Materials and methods. Control uninfected hepatocytes are shown in lane 1. Middle panel. E-2 immunoblots were quantified on a Kodak 4000 Imaging Station and software; P <0.05 for E-2 expression for primary (lanes 2–4) and secondary (lanes 5–7) infections when compared to control samples (lane 1). Results from triplicate samples of two independent experiments are shown. Lower panel. [³⁵S]-methionine labeling of HCV E-2 in the secondary infection of human hepatocytes infected with HCV genotypes 1, 2 or 3 (lanes 5–7) produced by the human hepatocyte primary infection. HCV infection was quantified by determining the radioactivity of immunopurified HCV E-2 (P<0.01); background radioactivity was negligible. β-Actin was used as a control for immunoprecipitation. Results from triplicate samples of three independent experiments are shown.