Ca²⁺ supplementation <i>in vitro</i> modulates cytokine production by primary human <i>i</i>NKT cells, Vδ2⁺ T cells, and MAIT cells.

PBMCs were isolated from the residual leukocyte units of healthy donors. PBMCs were stimulated for 4 h with 25 ng/ml PMA and 1 μg/ml ionomycin in either normal RPMI1640 medium (RPMInorm) or RPMI1640 medium supplemented with 1 mM Ca2+ (RPMIsuppl). (a) Human Vα24i NKT cells (live CD14- CD20- CD3+ 6B11+ cells) were analysed for the expression of TNF. (b, c) Human Vδ2+ T cells (live CD14- CD20- CD3+ γδTCRlow or Vδ2+ cells) were analysed for the expression of (b) TNF and (c) IFNγ. (d-f) Human MAIT cells (live CD14- CD20- CD3+ Vα7.2+ CD161+ cells) were analysed for the production of (d) IL-2, (e) TNF, and (f) IFNγ. Summary graphs (left panels) and representative data (right panels) from gated iNKT cells, Vδ2+ T cells, and MAIT cells are shown, respectively. Data were pooled from three (iNKT cells, Vδ2+ T cells; n = 9) and four independent (MAIT cells; n = 12) experiments with three samples each.