CD2-mediated IL-12–dependent signals render human γδ-T cells resistant to mitogen-induced apoptosis: analysis by 4-color flow cytometry.

PBMC cultures were initiated as described above. Those receiving day 0 signals (IFN-γ, IL-12, and anti-CD2 mAb S5.2) by convention were defined as protected. Those receiving no anti-CD2 mAb S5.2 or IL-12 on day 0 (PBS only) were defined as unprotected. All cultures received OKT3 and IL-2 24 hours later (day 1 mitogenic signals). Both αβ- and γδ–T-cell populations were first delineated by electronic gating on the corresponding αβ- and γδ-T cells defined by anti-CD3-APC and anti-TCR-γδ–PE mAbs. Apoptosis occurring in αβ- and γδ–T-cell populations was then determined examining the uptake of annexin V-FITC and PI in the respective gated events. Cells incubated with anti-human CD95/Fas mAb CH11 (mouse IgM,) or mouse IgM isotype control antibody were used as positive and negative controls, respectively, to define apoptotic, viable, and necrotic quadrants within dot plots. Percentages of αβ- or γδ-T cells appearing in the corresponding dot plot quadrants are indicated: viable (annexin⁻/PI⁻), apoptotic (annexin⁺/PI⁻), and necrotic (annexin⁺/PI⁺). Results shown are representative of experiments performed using materials obtained from at least 4 different persons.