FIG. 6.

Detection of nuclease-resistant HCV-RNAs in supernatants of various cell lines. Supernatants from parental Huh-7 cells (A), cell line 9-13 harboring a subgenomic replicon (B), and cell line 21-5 with an sfl genome (C) were harvested 6 days postseeding, cleared by low-speed centrifugation, filtered through 0.45-μm-pore-size filters, and pelleted by high-speed ultracentrifugation. Pellets were resuspended in PBS supplemented with carrier RNA, and aliquots were incubated for 4 h at room temperature with RNase A at the concentrations given below the bars. Some aliquots were also supplemented with 0.5% NP-40 (indicated by + [−, no NP-40 added]). HCV RNA was measured by quantitative RT-PCR performed in duplicate. Mean values of duplicates representing the number of HCV RNA molecules per milliliter of supernatant are given, and the error bars indicate the standard errors of the means.