The core dimer is formed via a bond between cysteine residues at amino acid position 128. (a) Site-directed mutagenesis of JFH1E2FL. (b) Immunoblot analysis of the core protein in MMFs collected from HuH-7 cells under nonreducing conditions 3 days post-transfection with JFH1E2FL (WT), JFH1C128A (C128A), JFH1C184A (C184A), or JFH1C128/184A (C128/184A) RNA. (c) Infectivity of culture medium collected and concentrated on day 5 posttransfection from HuH-7 cells transfected with WT, C128A, C184A, or C128/184A RNA. (d) Real-time qRT-PCR analysis of HCV RNA titers in culture medium collected at the indicated time points from HuH-7 cells transfected with WT, C128A, C184A, C128/184A, or PP/AA (JFH1PP/AA) RNA. (e) ELISAs of core protein levels in culture medium collected at the indicated time points from HuH-7 cells transfected with WT or C128A RNA. Data are representative of those from three independent experiments (b and c) or are the means ± standard deviations from three independent experiments (d and e).