The role of self-peptides in positive selection of CD4+ T cells has been controversial. We show that some self-peptides are presented by the MHC class II molecule I-A(b) in mice lacking Ii or H-2M but not in mice expressing a transgene-encoded peptide fused to I-A(b). In experiments using specific antibodies to block selection, these low-abundance self-peptides were implicated in the positive selection of some CD4+ T cells in H-2M-/- mice. However, all three mutant backgrounds failed to positively select two class II-restricted transgenic T cell receptors. Our findings suggest that minor components of the self-peptide repertoire can contribute to positive selection of a significant number of CD4+ T cells. In addition, the data suggest that T cell receptor repertoires selected in wild-type mice and in mice displaying limited spectra of self-peptides are distinct.