Oral keratinocytes trans infect HIV-1 to permissive PBMCs. TERT-2 or TE monolayers were inoculated and incubated for 6 h with lab-adapted HIV-1, IIIb or BaL. Tonsils were obtained from six donors (tissues 144, 193, 195, 196, 1101, and 223). Cells from each donor were propagated separately and TE cells were cultured as described in Materials and Methods. After incubation, cells were trypsinized, washed to remove non-internalized particles, and then co-cultured with PHA-activated PBMCs (2 × 105 cells) in PBMC growth media. To estimate HIV-1 trans infection from keratinocytes, PBMCs supernatants were collected on day 9 post inoculation and p24gag expression was estimated using ELISA. (A) TE cells from each donor differentially trans infect HIV-1 to PBMCs. (B) TERT-2 and TE 223 cells were tested side-by-side in the same experiments to compare HIV uptake and transfer. Mouse fibroblast cells (NIH 3T3) were included as a negative control. (C) To investigate the rate of HIV-1 trans infection over time, TERT-2 cells were trypsinized and washed to remove extracellular HIV-1 at indicated times post inoculation. TERT-2 cells from each time point were then co-cultured with PBMCs and p24gag production was analyzed. TERT-2 cells incubated with media only (no virus; NV) or heat-inactivated HIV-1 BaL (HV) were included as negative controls. Data in panel A represent the mean ± standard deviation of triplicate determinations in one experiment since the availability of primary tonsil cells from each donor was limited. Data in panel B and C are reported as the mean ± standard deviation from three independent experiments each performed in triplicate.