Two hundred twenty-four patients with leukemia transplanted with an unrelated donor between 1991 and 2003 at the Karolinska University Hospital were analyzed according to association between graft failure and ABO, RhD, MNSs, and Kidd blood group antigen compatibility. Median age was 29 years (range: 0-55). Conditioning consisted of total-body irradiation or busulfan-based myeloablative conditioning. A bone marrow graft was given to 152 patients, and 72 patients received peripheral blood stem cells. Most patients received graft-versus-host disease prophylaxis with cyclosporine and MTX. Graft failure (GF) was seen in 6 (2.7%) patients. In the multivariate analysis major ABO mismatch (odds ratio [OR] 14.9, 95% confidence interval [CI] 2.01-110, P = .008) and HLA-allele mismatch (6.42, 1.19-34.8, P = .03) was significantly associated to GF. In patients with and without major ABO mismatch the incidence of GF was 7.5% and 0.6% (P = .02), respectively. Using an ABO major mismatched graft increases the risk for GF after unrelated donor hematopoietic stem cell transplantation.