We have analyzed the ability of human gamma+/delta+ T cells to recognize a nominal antigen in association with MHC molecules. A TT-specific T cell line with approximately 40% gamma+/delta+ T cells was established from a hyperimmunized donor, D.F., by stimulation with antigen and autologous APC. Three DF-derived gamma+/delta+ clones were CD8+ as determined by immunofluorescence staining, and by Southern and Northern blotting with probes detecting delta chain rearrangement and delta and gamma chain transcripts, respectively. The gamma+/delta+ clones responded to stimulation with TT, but not TNP-BSA, and autologous APC by proliferation and IFN-gamma production. No proliferation or IFN-gamma production was detected when TT-specific T cell clones were stimulated with either TT or autologous APC only. The response to TT was enhanced by addition of exogenous IL-2. The use of allogeneic APC from 19 donors sharing one HLA-determinant with the autologous donor D.F., showed that the gamma+/delta+ T cells responded to TT with HLA-DR4-related restriction as measured by proliferation and IFN-gamma production. These results demonstrate that gamma/delta receptors can recognize non-MHC-encoded foreign antigen in a self-MHC-restricted fashion.