Fig. 3.

Infectivity of encapsidated, full-length HPV16 DNA. Virions encapsidating the HPV16 W12 genome were produced as in Fig. 2C and inoculated onto HaCaT cells. At 48 h after inoculation, cells were harvested, total RNA was isolated, and a single round of RT-PCR (A) or RT-PCR, followed by a second round of nested PCR (B and C), was performed with PCR primers (see Table 1) to detect E1 E4 spliced mRNAs. We amplified β-actin mRNA simultaneously as an internal standard. (A) Cells were inoculated with 100 to 1.6 vge per cell as indicated and assayed for infection by single round RT-PCR. (B) As indicated, HaCaT cells were mock-inoculated or inoculated with 100 vge per cell of virions that were untreated or treated with high-pH carbonate buffer for 24 h at 4°C or with unencapsidated, full-length HPV16 DNA. Total RNA extracted from HPV16-positive W12E cells was used as a positive control. (C) Before inoculating at 100 vge per cell, virions were incubated for 1 h at 4°C with 1:100 dilutions of HPV16 L1-neutralizing antibodies H16.7E or H16.V5. A nonneutralizing, anti-HPV16L1 antibody and an anti-HIV Gag antibody were used as isotypic negative controls.