Xenograft rejection in the discordant pig-to-primate model is dependent on binding of natural antibodies to gal-alpha [1-3]-gal epitopes on the porcine endothelial cell (EC). This leads to complement activation and deposition of activation products onto the membrane and results in perturbation of EC function and thrombus formation. Here we investigated the ability of human complement activation products to directly induce activation of porcine EC, with subsequent upregulation of adhesion and pro-coagulant molecules. Porcine aortic EC were isolated from wild-type and hDAF-transgenic pigs and incubated with human serum, either in the presence or absence of the soluble complement inhibitor TP10 (sCR1). Recombinant C5a, C1q-IgG immune complexes, C6-deficient human serum and serum containing anti-C9 Ab were used to identify EC activating complement products. Heat-inactivated human serum was used as a negative control. Cells were stained with antibodies against human C3, the MAC or with antibodies cross-reactive for porcine E-Selectin, VCAM-1 or Tissue Factor, and analyzed by flow cytometry. We found upregulation of E-Selectin and Tissue Factor on wild-type EC after incubation with human serum. This effect coincided with the deposition of C3 and MAC on the membrane of these cells. The addition of TP10 inhibited EC activation by up to 95%. In contrast, greatly reduced C3 and MAC deposition was detected on hDAF transgenic cells, and no complement-mediated EC activation was seen. Experiments with C6-deficient serum and incubation with anti-C9 Ab indicate a major role of the MAC in serum-mediated EC activation, whereas neither C5a nor C1q-IgG caused activation of EC. These data provide further explanation of the protective role of human DAF in the pig-to-primate xenotransplantation model.