Recognition of the class Ib antigen Qa-1 by a portion of alloreactive cytotoxic T lymphocyte (CTL) clones requires that the target cell express a second gene, termed Qa-1 determinant modifier (Qdm). We show that Qdm is identical to most D allele genes, excepting Dk, and that a nonamer peptide derived from D alloantigens restores CTL recognition on cells that lack the Qdm-encoded determinant. The equivalent Dk peptide has an Ala-->Val interchange at P3 and requires approximately 4 logs more peptide than the AlaP3 peptide for target cell lysis. Two of five CTL clones, not dependent on Qdm for target cell recognition, also recognize the Qdm peptide as well as the ValP3 variant. Although the Qdm peptide spans residues 3-11 from the leader, it requires the Tap transporters for its expression. Thus, the response against this class Ib molecule provides a tool for dissecting alloreactivity as well as pathways for antigen presentation.