Characterization of porcine platelet glycoproteins recognized by human natural "anti-gal" antibodies.
Human natural "anti-Gal" antibodies are specifically directed to Gal alpha 1-3Gal beta 1-4GlcNAc residues expressed on non-primate mammal and new world monkey cells. We investigated the relative involvement of purified IgG and IgM anti-Gal as xenoreactive natural antibodies (XNA). IgG and IgM were isolated from human plasma, and anti-Gal antibodies were purified by affinity chromatography on a Synsorb-14 column (Chembiomed, Edmonton, Alberta, Canada). Anti-Gal of both IgM and IgG classes represent the bulk of human XNA that bind to porcine platelets in enzyme-linked immunosorbent assay (ELISA). On immunoblots, normal human sera, as well as purified IgM and IgG fractions, reacted with 115-, 125-, 135-, 150-, 180-, 210-, and 240-kd) pig platelet proteins, whereas purified anti-Gal antibodies of both IgM and IgG classes mainly bound to 135-, 150-, 180-, and 210-kD glycoproteins. A low reactivity was observed in ELISA with anti-Gal free IgM and IgG, indicating that xenoantibodies are not solely directed to galactosyl epitopes. These antibodies revealed bands of 115, 125, and 240 kD, alpha-Galactosidase treatment of porcine platelet glycoproteins (gps) enriched by affinity chromatography abrogated the reactivity of 135- and 210-kD proteins. N- and O-glycosidase treatments demonstrated that alpha-galactosyl residues are located on the O-glycans of the 135-kD component. Finally, glycoproteins of 90 and 135 kD were identified by amino acid sequencing as the pig analogs of the human glycoproteins IIIa and IIb, respectively, whereas the 240-kD) component was identified as the porcine fibrinogen, using a new murine monoclonal antibody (naM147-7B6; IgG1) specific for its beta-chain.
Thibaudeau
K
K
Centre Régional de Transfusion Sanguine, Nantes, France.
Borche
L
L
Soulillou
J P
JP
Blanchard
D
D
D000595:Amino Acid Sequence; D000818:Animals; D000910:Antibodies, Heterophile / Q000276:immunology*; D000911:Antibodies, Monoclonal / Q000276:immunology; D015478:Antigens, Heterophile / Q000737:chemistry / Q000276:immunology*; D002240:Carbohydrate Sequence; D002846:Chromatography, Affinity; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000276:immunology*; D006001:Glycoconjugates / Q000276:immunology*; D006026:Glycoside Hydrolases / Q000494:pharmacology; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D010980:Platelet Membrane Glycoproteins / Q000737:chemistry / Q000276:immunology* / Q000302:isolation & purification; D011323:Primates / Q000276:immunology; D013045:Species Specificity; D013552:Swine / Q000276:immunology*
D000910:Antibodies, Heterophile; D000911:Antibodies, Monoclonal; D015478:Antigens, Heterophile; D000939:Epitopes; D006001:Glycoconjugates; D010980:Platelet Membrane Glycoproteins; D006026:Glycoside Hydrolases