baCD3/CD28-TK⁺ human lymphocytes lack lytic granules but strongly up-regulate CD40L on restimulation. At day 10, aCD3- or baCD3/CD28-TK⁺ lymphocytes were compared with the corresponding PBLs for CD40L up-regulation and for expression of lytic granules. (A) Before (top dot plots) and 2 hours after stimulation with PMA and ionomycin (bottom dot plots), cells were analyzed by flow cytometry for CD40L and CD8 expression after gating for CD3 in the case of PBLs and for LNGFR in the case of TK⁺ cells. Quadrants and percentages were set according to isotype control staining. The inserts report the percentages of cells for each quadrant. Results with cells from one representative donor of 6 are shown. Curves depict averages ± SD of the proportion CD4⁺ cells in PBLs or aCD3- or baCD3/CD28-TK⁺ lymphocytes that also express CD40L over time. (B) Cells were also analyzed by flow cytometry for the expression of granzyme A (Gr-A) and perforin B (Pf-B). Quadrants and percentages were set according to isotype control staining. The inserts report the percentages of cells for each quadrant. Results with cells from one representative donor of 3 are shown. Averages ± SD of the relative distribution of GrA^-PfB^-, GrA⁺PfB^-, or GrA⁺PfB⁺ in CD8⁺ cells from PBLs or aCD3- or baCD3/CD28-TK⁺ lymphocytes are reported (histograms). Symbols overlying aCD3- or baCD3/CD8-TK⁺–related bars are set for statistical comparison with the corresponding PBL-related symbols or bars (^*P < .05; ^**P < .01; ^***P < .005).