Capture of anti-(Galalpha1-3Gal) antibodies by immobilized Galalpha1-3Gal oligomers derived from carrageenan.
An immobilized Galalpha1-3Gal-bearing affinity ligand was prepared as a possible prophylactic binding site of human anti-(Galalpha1-3Gal) antibodies for the prevention of hyperacute rejection of pig xenografts. lambda-Carrageenan, a natural polysaccharide known to possess alternately alpha1-3 and beta1-4-linked D-galactopyranose sulphate residues, was selectively degraded by acetolysis and subsequently deacetylated to obtain lambda-carrageenan oligosaccharides. The resulting syrup was ultrafiltered to remove higher polymers and chromatographed on Sephadex G-15 to produce a series of sized oligosaccharide fractions. An immuno-dot-blot method was established on the basis of the competition between the lambda-carrageenan oligosaccharides in solution and bovine thyroglobulin (BTG), a glycoprotein known to express the Galalpha1-3Gal marker, as the solid-phase antigen. Increasing amounts of lambda-carrageenan fractions added to normal human plasma resulted in a progressive decrease in Galalpha1-3Gal-specific human IgM and IgG binding to BTG. Complete inhibition of Galalpha1-3Gal-specific human immunoglobulin was attained at concentrations less than 0.6 mg of carbohydrate/ml of human plasma with the more active fractions. These bioactive lambda-carrageenan oligosaccharides were immobilized on hydrazide-modified microporous nylon membranes, yielding capacities between 2.18 and 2.86 mg/ml of membrane. A subsequent decrease in the human anti-(Galalpha1-3Gal) antibody level in normal human plasma was demonstrated and proved with the established immuno-dot-blot procedure.
Klein
E
E
Kidney Disease Program, University of Louisville, Louisville, KY 40202, USA.
Euler
M
M
Vercellotti
J
J
D000818:Animals; D000906:Antibodies / Q000097:blood*; D000918:Antibody Specificity; D002240:Carbohydrate Sequence; D002351:Carrageenan / Q000737:chemistry*; D002850:Chromatography, Gel; D004187:Disaccharides / Q000276:immunology*; D005690:Galactose / Q000276:immunology*; D006801:Humans; D008024:Ligands; D008969:Molecular Sequence Data; D013552:Swine; D014183:Transplantation, Heterologous; D014312:Trisaccharides / Q000276:immunology
D000906:Antibodies; D004187:Disaccharides; D008024:Ligands; D014312:Trisaccharides; D002351:Carrageenan; D005690:Galactose