FIG. 2.

Sensitivity and specificity of LP-PCR and comparison with a nested Alu PCR protocol. (A to C) Viral DNA accumulation following cell-free infection in the presence or absence of inhibitors. HuT-78 T cells were infected using the centrifugal enhancement protocol at 0.5 TCID₅₀ per cell and cellular DNA prepared from infected cells harvested at 26 h p.i. 3TC and L-731,988 were used as specific inhibitors of reverse transcription and integration, respectively. (A) Sensitivity of LP-PCR (as measured by amplification of the HA8 integrated HIV DNA standard) and integrated HIV DNA accumulation following infection as measured by LP-PCR performed on 100 cell equivalents of Hirt pellet (chromosomal) DNA preparations. (B) Total reverse-transcribed DNA as measured by GAG-PCR performed on combined Hirt supernatant (extrachromosomal) and Hirt pellet (chromosomal) DNA samples. (C) Graphical representation of the accumulation of integrated HIV DNA. Data were obtained by PhosphorImager analysis of the bands in panel A. (D) Comparison of PCR detection of integrated HIV DNA by LP-PCR and Alu PCR. Chromosomal DNA was isolated from ACH-2 or 8E5 cells and shown to contain equivalent amounts of total HIV DNA by GAG-PCR (314-bp band). Sizes of expected bands for LP-PCR (measuring integrated HIV DNA) are given on the left (104-bp fragment), while the expected size of the product obtained following Alu PCR (also measuring integrated HIV DNA) is indicated on the right (351-bp fragment).