Responder cells [C57BL/6J X A.TL)F1 lymph node cells depleted of bursa equivalent-derived (B) cells by filtration through nylon wool columns] were activated against incompatible K-region and I-region products together under conditions where these antigens are presented on separate stimulator cells. The resulting T blasts were stained with different concentrations of antisera directed against incompatible stimulator K-region or I-region products, or both. We obtained results that strongly suggest that in these cultures each activated responder blast stains with antiserum directed against either K-region or I-region products, but not both. Responder blasts from the same cultures were treated with antiserum and complement (C) directed against either Ly-1.2 or Ly-2.2 T-cell-specific surface antigens. Anti-Ly-1.2 serum and C specifically eliminates virtually all responder blasts staining with antiserum directed against stimulator I-region products; whereas anti-Ly-2.2 serum reduces to background levels the proportion of cells staining with antiserum against stimulator K-region products. The results obtained suggest that T cells binding stimulator K-region and I-region products, respectively, belong to two different subclasses distinguishable by their Ly phenotypes. Possible explanations for this association of T- cell subclass and specificity are discussed.