FIG. 3.

HCV released from cells transfected with Pol I-driven plasmids. (A) HCV particle secretion from the transfected cells. The culture supernatant of Huh-7 cells transfected with Pol I-driven plasmids containing wild-type or mutated HCV genome were harvested on days 2, 4, and 6 and assayed for HCV core protein levels. The data for each experiment are averages of triplicate values with error bars showing standard deviations. (B) Sucrose density gradient analysis of the culture supernatant of pHHJFH1-transfected cells. Culture supernatant collected on day 5 p.t. was cleared by low-speed centrifugation, passed through a 0.45-μm-pore-size filter, and concentrated ∼30-fold by ultrafiltration. After fractionating by sucrose density gradient centrifugation, the core protein level and viral infectious titer of each fraction were measured. (C) Kinetics of core protein secretion from cells transfected with pHHJFH1 or with JFH-1 genomic RNA. A total of 10⁶ Huh-7 cells were transfected with 3 μg of pHHJFH1 or the same amount of in vitro-transcribed JFH-1 RNA by electroporation. The cells were passaged every 2 to 3 days before reaching confluence. Culture supernatant collected on the indicated days was used for core protein measurement. The level of secreted core protein (pmol/liter) is expressed on a logarithmic scale. The data for each experiment are averages of triplicate values with error bars showing standard deviations.