Figure 3

Polyfunctional analysis of Mtb PPD-specific cytokine expression in CD4 T-cells and NK cells 12 months post BCG vaccination of UK adolescents. A) Dot plots from one individual donor are shown comparing simultaneous expression of IFNγ (x-axis) and TNFα (y-axis). Previous gating was used to isolate singlet, live cells that were either CD3⁺CD4⁺ (CD4 T-cells) or CD3^-CD56⁺ (NK cells) as indicated for each plot. Responses in unstimulated and Mtb PPD-stimulated samples are shown. Values indicate the proportion (as a percentage) of the indicated subset present in that quadrant. B) Boolean gating and Spice analysis was used to determine the extent of polyfunctionality within the 12 month post-BCG vaccination group (n = 12). Bar charts represent, for either CD4 T-cells or NK cells, the median percentage (and interquartile range) of each subset expressing the indicated cytokine combination (x-axis labels). Pie segments represent the average proportion, across the group, of cells expressing the indicated cytokine combination (pie chart legend) within the total population of cells expressing either IFNγ or TNFα.