Natural and elicited antipig antibodies (Abs) lead to acute humoral xenograft rejection (AHXR). Ten baboons underwent heterotopic heart transplantation (Tx) from human decay-accelerating factor (hDAF) pigs. Depletion of anti-Galα1, 3Gal (Gal) Abs was achieved by the infusion of a Gal glycoconjugate from day - 1. Immunosuppression included induction of antithymocyte globulin, thymic irradiation, and cobra venom factor, and maintenance with a human antihuman CD154 mAb, mycophenolate mofetil, and methylprednisolone, heparin and prophylactic ganciclovir were also administered. Pig heart survival ranged from 4 to 139 (mean 37, median 27) days, with three functioning for >50 days. Graft failure (n = 8) was from classical AHXR , thrombotic microangiopathy , or intragraft thrombosis , with death (n = 2) from pneumonia , or possible drug toxicity (with features of thrombotic microangiopathy) . Anti-Gal Abs (in μg/mL) were depleted by Gal glycoconjugate before graft implantation from means of 41.3 to 6.3 (IgM) and 12.4-4.6 (IgG), respectively, and at graft excision were 6.3 and 1.7 μg/mL, respectively. No elicited Abs developed, and no cellular infiltration was seen. The treatment regimen was effective in maintaining a low anti-Gal Ab levels and in delaying or preventing AHXR. The combination of costimulatory blockade and heparin with Tx of a Gal-negative pig organ may prolong graft survival further.