A group of 42 renal transplants performed in the presence of a T-cell-positive crossmatch were analyzed to determine the class and specificity of the donor-reactive cytotoxic antibodies. Dithiothreitol (DTT) was used to reduce IgM antibodies and a monoclonal antibody directed at a monomorphic determinant present on all HLA class I antigens (PA2.6) was used to inhibit cytotoxicity of anti-HLA class I antibodies. Sera from 26 of the positive crossmatches were considered to be autoreactive, and the positive crossmatch proved to be due to IgM and not directed at HLA class I in each case. One year graft survival was 100% in the 5 living-related and 60% in the 21 cadaver donor transplants, of which 10 were regrafts. Of the 42 positive crossmatches, 16 were not due to autoantibody. One was positive in the current serum taken at the time of transplantation, and this graft was rejected hyperacutely, while 15 were positive with peak but not current serum samples. Of the positive crossmatches, 12 were inhibited by PA2.6 demonstrating that they were directed at HLA class I antigens. PA2.6 inhibition could not be shown in 3 and in 1 DTT reduction was technically unsatisfactory. While 4 of the 7 positive crossmatches due to IgM antibodies were successful, the 7 transplants performed with positive crossmatches due to IgG antibodies all failed. DTT reduction and inhibition of cytotoxicity by PA2.6 helps to define positive crossmatches with donor T cells that are not associated with graft failure. Transplantation in the presence of a peak positive T cell crossmatch due to an anti-HLA antibody might only be successful if the antibody in the peak serum is of the IgM class.