Identification of erythrocyte Gal alpha 1-3Gal glycosphingolipids with a mouse monoclonal antibody, Gal-13

The Gal alpha 1-3Gal structural determinant has been found to have a unique distribution in mammals. Although this determinant is abundantly expressed by erythrocytes and nucleated cells of many mammals, it has not been detected in human cells. However, our previous studies (Galili, U., Rachmilewitz, E. A., Peleg, A., and Flechner, I. (1984) J. Exp. Med. 160, 1519-1531; Galili, U., Clark, M. R., and Shohet, S. B. (1986) J. Clin. Invest. 77, 27-33) have suggested that this epitope is present in small amounts and may be involved in immune-mediated destruction of senescent human erythrocytes. To have a means for exploring this possibility and for studying the species and tissue distribution of this epitope we have raised a monoclonal antibody (Gal-13) which specifically binds to glycoconjugates with a nonreducing terminal Gal alpha 1-3Gal disaccharide. Mice were immunized with rabbit erythrocytes, which express an abundance of glycoconjugates with Gal alpha 1-3Gal epitopes. Clones were screened with a solid-phase binding assay (enzyme-linked immunosorbent assay) for antibodies which bound to ceramide pentahexoside (Gal alpha 1-3Gal beta 1-4GlcNAc beta 1-3-Gal beta Gal beta 1-4Glc1-1Cer) but not to ceramide trihexoside (Gal alpha 1-4Gal beta 1-4Glc1-1Cer). Gal-13 bound to a number of neutral glycosphingolipids from rabbit and bovine erythrocytes. These glycosphingolipids have previously been shown to be a family of linear and branched polylactosamine structures, which have non-reducing terminal Gal alpha 1-3Gal epitopes. The antibody did not bind to the human blood group B glycolipid, Gal alpha 1-3(Fuc alpha 1-2)Gal beta 1-4GlcNAc beta 1-3Gal beta 1-4Glc1-1Cer, and, therefore, branching at the penultimate galactose blocks Gal-13 binding. However, after removal of the fucose from the B antigen Gal-13 recognized the resulting derivative. Other Gal alpha 1-3Gal glycosphingolipids with an isogloboside or globoside core structure were not recognized by Gal-13 suggesting that the antibody binds to Gal alpha 1-3Gal carried by a lactosamine core structure. Gal-13 has been used to demonstrate that the Gal alpha 1-3Gal ceramide pentahexoside has been evolutionarily conserved in red cells of animals up to the stage of New World monkeys but is not found in Old World monkey red cells.(ABSTRACT TRUNCATED AT 400 WORDS)