Purified peripheral blood gamma delta T cells proliferated vigorously in response to killed Mycobacterium tuberculosis (M. tb.) in the presence of PBMC but not in the presence of T cell-depleted (E-) feeder cells. Addition of graded numbers of autologous CD4 T cells to E- feeder cells reconstituted in a dose-dependent fashion the response of V gamma 9-expressing gamma delta T cells to M. tb. IL-2 was identified as the major CD4 T cell-derived helper factor required for gamma delta T cell proliferation after stimulation with M. tb. In addition, neutralizing anti-IFN-gamma but not anti-IFN-alpha Ab inhibited the responsiveness of V gamma 9 T cells, suggesting that endogenously produced IFN-gamma was involved in the activation of gamma delta T cells by M. tb. Although gamma delta T cells could not proliferate on their own in the absence of CD4 T cells (or exogenous IL-2), the appearance of IL-2 receptors (CD25) was triggered in the absence of CD4 T cells. Furthermore, IL-10 strongly inhibited the activation of V gamma 9 T cells among unfractionated PBMC responder cells. Similarly, the responsiveness of purified gamma delta T cells to M. tb. occurring in the presence of CD4 T cells was strongly inhibited by IL-10, whereas the activation occurring in the presence of exogenous IL-2 was not impaired. These results show that interactions with Th1-type CD4 T cells are required for efficient activation of peripheral blood gamma delta T cells by M. tb. In addition, our results have practical implications for creating experimental conditions aimed at identifying V gamma 9-selective (myco)bacterial ligands.