The recent development of a reliable murine T [thymus-derived] lymphocyte proliferation assay has facilitated the study of T lymphocyte function in vitro. In this paper, the effect of anti-histocompatibility antisera on the proliferative response was investigated. The continuous presence of anti-Ia [immune response associated] antisera in the cultures inhibited the responses to the antigens poly (Glu58Lys38Tyr4) [GLT], poly (Tyr, Glu) poly D,L Ala.sbd.poly Lys [T,G)-A.sbd.L], poly (Phe, Glu)-poly D,L Ala.sbd.poly Lys [(.vphi. G)-A.sbd.L], lactate dehydrogenase H4, staphylococcal nuclease and the IgA [immunoglobulin A] myeloma protein, TEPC 15. The T lymphocyte proliferative responses to all of these antigens were previously shown to be under the genetic control of major histocompatibility-linked immune response genes. The anti-Ia antisera were also capable of inhibiting proliferative responses to antigens such as PPD [purified protein derivative], to which all strains respond. Antisera directed solely against H-2K or H-2D antigens did not give significant inhibition. Anti-Ia antisera capable of reacting with antigens coded for by genetically defined subregions of the I locus were capable of completely inhibiting the proliferative response. In the 2 cases studied, GLT and (T,G)-A.sbd.L, an Ir [immune response] gene controlling the T lymphocyte proliferative response to the antigen was previously mapped to the same subregion as that which coded for the Ia antigens recognized by the blocking antisera. In F1 hybrids between responder and nonresponder strains, the anti-Ia antisera showed haplotype-specific inhibition. That is, anti-Ia antisera directed against the responder haplotype could completely block the antigen response controlled by Ir genes of that haplotype; anti-Ia antisera directed against Ia antigens of the nonresponder haplotype gave only partial or no inhibition. Since this selective inhibition was reciprocal depending on which antigen was used, the mechanism of anti-Ia antisera inhibition was probably not cell killing or a nonspecific turning off of the cell but rather a blockade of antigen stimulation at the cell surface. The selective inhibition demonstrates a phenotypic linkage between Ir gene products and Ia antigens at the cell surface. These results, coupled with the known genetic linkage of Ir genes and the genes coding for Ia antigens, suggest that Ia antigens are determinants on Ir gene products.