Figure 1.

Time-of-addition experiments using CM-II-sPLA₂ against HCV and DENV. (a) Immunoblotting analysis. The HCV NS3 (upper panel) and DENV PrM (lower panel) expression levels were examined by immunoblotting. (i) Pretreatment of the cells: Huh7it-1 cells were treated with decreasing concentrations of CM-II-sPLA₂ (1,000, 100, 10 and 1 ng/ml) for 1 h. Then, the cells were inoculated with HCV or DENV in the absence of CM-II-sPLA₂ for another 1 h and cultured for 24 h in the absence of CM-II-sPLA₂. (ii) Pretreatment of the virus: HCV and DENV were incubated with CM-II-sPLA₂ for 1 h, and the mixtures were inoculated onto Huh7it-1 cells. After 1 h, the cells were cultured for 24 h in the absence of CM-II-sPLA₂. (iii) Post-entry treatment: Huh7it-1 cells were inoculated with HCV or DENV in the absence of CM-II-sPLA₂. After 1 h, the cells were cultured for 24 h in the presence of CM-II-sPLA₂. (–), Untreated control. The full-length gels and blots are shown in Supplementary Figs S2 and S3. (b) qRT-PCR analysis. The HCV RNA (upper panel) and DENV RNA (lower panel) levels in cells prepared under the same conditions described in (a) were quantified by qRT-PCR. (–), Untreated control. Data are presented as the percentage of the untreated control.