To test directly the determinant selection hypothesis of immune response gene function, we primed strain 13 T lymphocytes in vitro with allogeneic bovine insulin pulsed strain 2 macrophages. Strain 2 macrophages were found to be fully competent to present bovine insulin B chain to strain 13 T cells despite the fact that strain 2 guinea pigs are normally totally unresponsive to this antigen. These results are incompatible with a strict interpretation of the determinant selection hypothesis, which would have predicted that strain 2 macrophages would have been restricted to the presentation of A chain loop determinants. In addition, a comparison of the reactivity profiles of self-Ia- and allo-Ia-restricted strain 13 T cells to a series of synthetic B chain peptide fragments revealed that the allo-Ia-restricted populations could be activated by autologous guinea pig insulin. Taken together, these observations strongly suggest that the clonal deletion of self-reactive cells is likely to be I region restricted and that nonresponsiveness to any protein antigen may result from a restriction in the T cell repertoire that is generated during ontogeny by a clonal deletion mechanism of tolerance to self.