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Identification of porcine membrane antigens involved in the cytotoxic response mediated by human xenoreactive antibodies
Author(s) -
Aspeslet Launa J.,
Chackowsky Patricia,
Sekhon Harbuksh,
Malcolm Andrew J.,
Mosleh Zina,
Koshal Arvind,
Yatscoff Randall W.
Publication year - 1996
Publication title -
xenotransplantation
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.052
H-Index - 61
eISSN - 1399-3089
pISSN - 0908-665X
DOI - 10.1111/j.1399-3089.1996.tb00112.x
Subject(s) - antigen , cytotoxicity , antibody , western blot , blot , microbiology and biotechnology , cytotoxic t cell , biology , chemistry , immunology , biochemistry , in vitro , gene
The identification of xenoantigens on the surface of endothelial cells is important for understanding the mechanism of hyperacute rejection and development of abrogating methods. The objective of our study was to identify the porcine antigens that, when bound by xenoreactive antibodies in human serum, result in cytotoxicity of porcine cells. Human AB and O sera were adsorbed with porcine aortae, erythrocytes, platelets, and a broad spectrum of immobilized carbohydrate moieties (Synsorbs). Aortae and erythrocytes were able to adsorb the xenoreactive antibodies that were cytotoxic to porcine cells (LLC‐PK1), determined using an MTT cytotoxicity assay. Only carbohydrates having the αGal(1–3)βGal(1–4) moieties (Synsorbs 90, 115) were able to significantly reduce cytotoxicity with both types of sera. Western blots of porcine aortic endothelial cells (PAEC) and LLC‐PK1 cell membrane extracts probed with unadsorbed sera indicate the binding of xenoreactive IgM to approximately 17 and 11 antigen bands, respectively, having molecular weights ranging from 20–133 kDa. Anti‐IgG development showed 8 and 11 antigen bands on PAEC and LLC‐PK1 membrane preparations, respectively. When blots were performed using adsorbed AB sera, the binding to all antigens was still observed. When Synsorb 90 bound antibodies were used to probe the blots, the majority of antigens were still detected. This suggests that the binding of xenoantibodies to the most prominent antigens, as detected by Western blot procedures may not be the ones to which cytotoxic xenoreactive antibodies bind. Alternative approaches are required to identify such antigens.