Fig. 3.

BCG-, M. vaccae- and M. obuense-induced activation of γδ T-cells is dependent on CD4⁺ cells. a γδ T-cells were cultured overnight with PMA/I, heat-killed BCG, M. vaccae (Mv) and M. obuense (Mo) and CD69 expression measured. Representative flow cytometric histogram plots from one of two donors are shown. Expression for untreated cells is shown in grey fill. b Culture supernatants from γδ T-cells were screened for their cytokine content using cytometric bead arrays. Mean values for n = 3 are shown. c γδ T-cells were cultured with PHA/IL-2, BCG, Mv and Mo for 6 days and ³H incorporation measured for the last 16 h of culture. Mean counts per minute (CPM) are shown for n = 2. d PBMCs were sequentially depleted of CD14⁺, CD19⁺ and CD56⁺ cells and stimulated overnight with BCG, Mv and Mo prior to measuring CD69 expression on gated Vδ2⁺ cells. Data were standardised by subtracting untreated scores from test scores. Mean values for n = 3 are shown. e γδ T-cells were co-cultured overnight with CD56⁺ cells (top graph) or a combination of CD4⁺ and CD8⁺ cells (bottom graph) in the presence of mycobacteria prior to measuring IFN-γ expression on gated Vδ2⁺ cells. Mean values for n ≥ 2 are shown. f γδ T-cells were co-cultured overnight with either CD8⁺ (top graph) or CD4⁺ (bottom graph) in the presence of mycobacteria prior to measuring IFN-γ expression on gated Vδ2⁺ cells. Mean values for n = 3 are shown. Error bars represent SD