According to some models of T cell education, tolerance, and autoimmunity, recognition of MHC molecules by T cells may depend on the nature of the APC expressing the MHC/Ag complex. To examine this, a panel of 23 I-Ad-restricted, alloreactive T cells were used to probe MHC class II molecules expressed on established lines representing different lineages. Surprisingly, we observed cell type-specific reactivity in the majority of the T cells. In all, 18 different reactivity patterns were identified. The patterns observed suggests that MHC reactivity can be organized into a hierarchical pattern for both the APC and the T cells. Experiments assessing T cells' avidities for allogeneic targets, ability to produce lymphokines, and expression of accessory molecules revealed no predictive correlation with the hierarchical reactivity pattern, nor did experiments measuring allogeneic target cells' expression of known accessory molecules and ability to stimulate Ag-specific hybridomas. These results suggest that the differential reactivity cannot be accounted for by accessory molecule discrepancies among the APC, but rather might reflect deficiencies in the ability of the various APC to engage the Ag-specific T cell receptor. Collectively, these data indicate that a significant fraction of allorecognition of MHC class II is cell-type dependent and that cell-type-specific recognition may relate to peptide-specific recognition requirements by the Ag receptor of alloreactive T cells.