IFN-γ production of activated γδ T cells.

PBMC were incubated with pamidronate (40 μmol/L) or medium alone. For a determination of the kinetics of IFN-γ secretion, supernatants were collected at indicated time points, and cytokine concentration was measured by ELISA. Each bar represents the mean values in triplicate for 1 representative donor (A). Intracellular IFN-γ expression of γδ T cells in response to IPP or pamidronate was measured by single-cell analysis after PBMC culture in medium alone or in the presence of IPP (40 μmol/L) or pamidronate (40 μmol/L) for 72 hours. After surface staining with a γδ TCR mAb, cells were fixed, permeabilized, and intracellularly stained with mAb against IFN-γ. Percentages of IFN-γ⁺ γδ T cells are given in the upper right panels (B). Controls using an isotype-matched control mAb in the presence of medium alone, IPP, or pamidronate always revealed less than 10% positive γδ T cells (data not shown). Results shown are representative for 3 independent experiments with different healthy donors.