An Attempt to Raise Antibodies against 2-O-α-D-Galactopyranose, the Disaccharide Unit of Collagen

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Abstract
The p-isothiocyanatophenyl β-glycoside of 2-O-α-D-glucopyranosyl-D-galactopyranose, the disaccharide unit of collagen, was coupled to bovine serum albumin for use as an artificial antigen. The conjugate precipitated with concanavalin A but not with the Ricinus communis lectin. Rabbit antisera raised against the conjugate were characterized by immunodiffusion, quantitative precipitation, and hapten inhibition. After absorption with bovine serum albumin the antisera exhibited a specificity for the introduced p-isothiocyanatophenyl 2-O-α-D-glucopyranosyl-β-D-galactopyranoside. The relative importance of an α-configuration and the ordered sequence of the two sugars for binding to antibody were observed. The antibody-combining site was shown to involve the aglycone portion of the introduced hapten. 5-O-(2-O-α-D-glucopyranosyl-O-β-D-galactopyranosyl)-hydroxylysine and 5-O-(2-O-α-D-glucopyranosyl-O-β-D-galactopyranosyl)-hydroxylysine peptide, the glycopeptide unit in collagens, were poor inhibitors of the conjugate-antibody precipitation system. However, acetylation of the ε-amino groups of the hydroxylysyl residues converted these noninhibitory substrates into considerable inhibitors, suggesting that a charged -NH3+ group may interfere with binding of the haptenic groups to antibody.

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