Separation and partial sequence analysis of blood group A-active oligosaccharides by affinity chromatography using monoclonal antibodies.

An affinity column containing an anti-blood group A monoclonal antibody coupled to Sepharose beads specifically retards oligosaccharides with the nonreducing trisaccharide sequence GalNAc alpha 1-3(Fuc alpha 1-2)Gal beta 1-R. Three A-active oligosaccharides, A-tetra, A-penta, and A-hepta, elute as retarded peaks, well-separated from unbound sugars. A-hepta, which contains a difucosylated type 1 (Leb) core structure, elutes much later than A-tetra or A-penta and can be completely separated from the latter oligosaccharides by affinity chromatography. The order of elution of the oligosaccharides agrees with their previously determined specific molar activities as inhibitors of quantitative immune precipitation [H.-T. Chen, and E. A. Kabat, (1985) J. Biol. Chem. 260, 13208-13217]. Treatment of A-hepta with Charonia lampas alpha-galactosaminidase abolishes its binding by the anti-A affinity column and converts it to a Leb-active oligosaccharide (lacto-N-difucohexaose I) that is specifically retarded on a second affinity column containing an anti-Leb monoclonal antibody.