Acute antibody-mediated (humoral) rejection is a major cause of morbidity, graft loss, and mortality among heart transplant patients. Herein we have presented our experience using C4d to characterize humoral rejection.All nonformalin-fixed cardiac graft biopsies (protocol or emergency) received between May 2007 and May 2008 were examined by immunofluorescence for C4d.One hundred twelve endomyocardial biopsies from 25 transplanted patients included 20 males and 5 females of ages ranging from 3 to 71 years. The number of biopsies per subject varied from 1 to 11; the timespan between transplantation and the diagnostic biopsies ranged from days to 8 years. Thirteen biopsies showed acute humoral rejection (intramyocardial capillaries positive for C4d); 31, acute cellular rejection (grades 1R, 2R); 7, both humoral and cellular rejection; and 1, acute humoral rejection and allograft vasculopathy. Some of the positive biopsies belonged to the same person, and some to transplanted individuals with signs and symptoms suggestive of rejection, while others did not. The persistence of humoral rejection, despite the disappearance of a cellular component, correlated with slower clinicoechocardiographic improvement.C4d positivity is a morphologic sign of humoral rejection. It may hasten the appearance and/or worsening of allograft vasculopathy independent of patient age or posttransplantation time.