Proliferative response of human γδ T cells to LA in the presence of moDC. (A) The percentage of γδ T cells was determined by flow cytometry. Similar results were observed in other five healthy donors. (B) In 3H-TdR incorporation assay, purified γδ T cells (99% purified) were co-cultured with LA in the presence or absence of moDC. Results are shown as cpm (mean ± SD) of five experiments. * P < 0.05 vs. γδ T, γδ T+LA or γδ T+DC respectively, * *P < 0.05 vs. γδ T. (C) Irradiated moDCs were fixed before or after LA stimulation. The proliferation of γδ T cells was determined as described in Methods and Materials. *P > 0.05 vs nonfixed moDCs. (D) The reduction of fluorescence of CFSE labeled γδ T cell-enriched PBMCs was detected after 72 h by gating on TCRγδ+ population. The top row shows dot plots; the bottom row shows the overlaps of histograms corresponding to top dot plots, among which the left histograms are the overlaps of γδT (black line histogram), γδ T+LA (dark grey filled histogram) and γδ T+DC (light grey filled histogram), and the right histograms are the overlap of γδ T+LA (dark grey filled histogram) and γδ T+LA+DC (black filled histogram). All experiments were performed at least three times with consistent results. γδ T: γδ T cells; LA: lipid A; DC: monocyte derived dendritic cells (moDC), IPP: isopentenyl pyrophosphate. These abbreviations above also appear in Fig. 2, 3, 4 and 5. Non-F, P-F or F-P respectively represent the groups, in which non-fixed moDCs, moDCs pulsed with LA before fixation, or moDCs fixed before pulsed with LA were used (Fig. 2C).