Adipose tissue is an attractive source for somatic stem cell therapy. Currently, human adipose tissue-derived stromal cells/mesenchymal stem cells (hADSCs/MSCs) are cultured with fetal bovine serum (FBS). Recently, however, not only human embryonic stem cell lines cultured on mouse feeder cells but also bone marrow-derived human MSCs cultured with FBS were reported to express N-glycolylneuraminic acid (Neu5Gc) xenoantigen. Human serum contains high titers of natural preformed antibodies against Neu5Gc. We studied the presence of Neu5Gc on hADSCs/MSCs cultured with FBS and human immune response mediated by Neu5Gc. Our data indicated that hADSCs/MSCs cultured with FBS expressed Neu5Gc and that human natural preformed antibodies could bind to hADSCs/MSCs. However, hADSCs/MSCs express complement regulatory proteins such as CD46, CD55, and CD59 and are largely resistant to complement-mediated cytotoxicity. hADSCs/MSCs cultured with FBS could be injured by antibody-dependent cell-mediated cytotoxicity mechanism. Further, human monocyte-derived macrophages could phagocytose hADSCs/MSCs cultured with FBS and this phagocytic activity was increased in the presence of human serum. Culturing hADSCs/MSCs with heat-inactivated human serum for a week could markedly reduce Neu5Gc on hADSCs/MSCs and prevent immune responses mediated by Neu5Gc, such as binding of human natural preformed antibodies, antibody-dependent cell-mediated cytotoxicity, and phagocytosis. Adipogenic and osteogenic differentiation potentials of hADSCs/MSCs cultured with heat-inactivated human serum were not less than that of those cultured with FBS. For stem cell therapies based on hADSCs/MSCs, hADSCs/MSCs that presented Neu5Gc on their cell surfaces after exposure to FBS should be cleaned up to be rescued from xenogeneic rejection.