Time-course analysis of degranulation and cytokine responses by different T-cell and NK cell subsets from healthy adults. Freshly isolated, resting PBMCs from healthy adult volunteers were incubated alone or mixed with target cells and mAbs, as indicated, at 37°C. Thereafter, cells were surface stained with fluorochrome-conjugated antibodies to CD3, CD4, CD8, CD56, CD57, and CD107a, followed by fixation, permeabilization, and intracellular staining with antibodies to MIP-1β, TNF-α, IFN-γ, and CD69. (A) Time course of functional responses on CD8⁺ T-cell subsets and CD3⁻CD56⁺ NK cells after target cell stimulations, as indicated. Values represent the means ± SD of 6 donors from 1 of 3 independent experiments. (B) Indicated lymphocyte cells were gated and a Boolean gating strategy was used for analysis. Pie charts represent the frequency of cells positive for the given number of measured responses (ie, CD107a, MIP-1β, TNF-α, and IFN-γ). Therefore, cells can be categorized into the number of responses they display. Arcs depict the relative frequency of cells specifically positive for CD107a, MIP-1β, TNF-α, and/or IFN-γ staining, as indicated. Values represent the means of 6 different donors from 1 of 3 independent experiments.