Quantitative evaluation of porcine endothelial cell antigens recognized by human natural antibodies: An analysis by Western blotting
Xenoreactive natural antibodies recognize a series of glycoproteins in porcine endothelial cell membranes, the epitopes being Galα1-3Gal substitutions on those proteins. We recently identified the glycoprotein antigens as members of the integrin family, von Willebrand factor, and DM-GRASP. The antibodies that react with these structures are adsorbed during perfusion of a porcine organ, indicating that these antigens may be the biologically relevant targets. To evaluate the relative importance of the glycoproteins as targets of xenoreactive natural antibodies, immunoreactive electrophoretic bands containing these glycoproteins were analyzed by photodensitometry. The relevance of these antigens in the recognition of endothelial cell antigens by xenoreactive natural antibodies was assessed by an analysis of the differences between antigens recognized by normal human serum and those recognized by human serum that had been depleted of xenoreactive natural antibodies. Depletion of xenoreactive natural antibodies was performed by adsorption of serum on cultured porcine aortic endothelial cell monolayers and by perfusion through a porcine kidney. The analysis revealed that greater than 91% of the antibodies binding to porcine cell surfaces are specific for antigens identified in porcine endothelial cell membrane extracts. Quantitative analysis of antigens recognized by affinity purified antibodies confirmed that integrins are indeed the primary xenogeneic targets recognized on Western blots of porcine aortic endothelial cell membranes. © Munksgaard, Copenhagen.
Lin, SS; Parker, W; Holzknecht, ZE; Platt, JL
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