This report describes the kinetics of T-cell responses to a panel of mycobacterial antigens (PPD-M, PPD-A, ESAT-6, Ag85, 38kD, MPB64, MPB70, MPB83, hsp 16.1, hsp65, and hsp70) following experimental infection of cattle with Mycobacterium bovis. Increased antigen-specific lymphocyte proliferation, gamma interferon, and interleukin-2 responses were observed in all calves following infection. Positive lymphocyte proliferation and cytokine responses to PPD-M and ESAT-6 were observed throughout the infection period studied. In contrast, responses to all other antigens were more variable and were not constantly present, suggesting that antigen cocktails rather than individual antigens should be used for immunodiagnosis. The detection of cytokine responses in the absence of lymphocyte proliferation, particularly during the early stages of infection, suggests a role for antigen-specific cytokine readout systems in the early identification of M. bovis infection in cattle.