A limiting dilution assay was used to compare the frequency of cytotoxic T lymphocyte precursors (CTL-P) which respond to H-2 antigens presented with additional background differences, with the frequency of CTL-P which respond to H-2 antigens on a self background. Individual cultures were divided and assayed for cytotoxic activity on the two targets sharing H-2, but not the background; no cultures were seen which clearly killed one and not the other, and the same frequency of CTL-P was measured on target cells that differed from the responder only at H-2 and on target cells that differed also in the background, irrespective of the background of the stimulator. Thus, the assumption that allospecific cytotoxic T lymphocytes (CTL) recognize H-2 plus minor histocompatibility antigens dose not serve as an adequate explanation for the high frequency of allospecific CTL. The data also suggest that the two allelic forms of beta2-microglobulin do not contribute to the alloantigenic determinant.