BACKGROUND The association between HLA class II antigens and the production of red cell autoantibodies was investigated in this study. METHODS Thirty-one individuals with a positive direct antiglobulin test (DAT) and 85. DAT-negative controls (cadaveric organ donors) were typed using a DNA-based method for class II HLA typing, that is, the polymerase chain reaction with sequence-specific primers. Amplified HLA alleles corresponding to the serologic specificities DR 1-18 and DQ 1-9 were identified. RESULTS Analysis of DR and DQ frequencies showed that HLA-DQ6 was less frequent in DAT-positive individuals (19% vs. 53% in the control population), with a p value of 0.0014 and a corrected p value of 0.059, and relative risk of 0.23 (95% Cl = 0.09-0.58). The frequency of HLA-DQ6 was higher in asymptomatic DAT-positive blood donors (n = 8, 38% DQ6 positive, p = 0.48) than in DAT-positive hospital and clinic patients (n = 23, 13% DQ6 positive, corrected p value = 0.030, relative risk = 0.13, 95% Cl = 0.04-0.41), 96 percent of whom had evidence of clinical hemolysis. CONCLUSIONS This study demonstrates that HLA-DQ6 may have a negative association with a positive DAT result in patients with evidence for hemolysis, and may be a resistance antigen for clinically relevant red cell autoantibodies.