AFFINITY-PURIFIED ANTIBODIES AGAINST ALPHA-GALACTOSYL RESIDUES FROM HUMAN SERUM - COMPARISON OF THEIR BINDING IN BOVINE TESTICULAR TISSUE WITH THAT OF THE GRIFFONIA-SIMPLICIFOLIA LECTIN (GSI-B-4) AND IMPACT OFLABELING ON EPITOPE LOCALIZATION

alpha-Galactosyl residues in the carbohydrate part of cellular glycoco njugates can serve as cell type-associated markers and are implicated in intercellular adhesion and biosignaling. This biological significan ce explains the interest to characterize probes with respective specif icity as the Griffonia simplicifolia I-isolectin B-4. Due to the docum ented occurrence of an alpha-galactoside-binding immunoglobulin G frac tion in human serum we compared the extent of binding and its pattern for the lectin and the antibody using surface-immobilized extract prot eins and fixed sections of bovine testicular tissue with known lectin reactivity. The antibody fractions were obtained either solely from af finity chromatography isolation on immobilized melibiose or after an a dditional step to deplete this fraction of galactoside-binding activit ies without pronounced specificity to the alpha-anomeric linkage. They yielded a rather indistinguishable reactivity in comparison to that o f the lectin, when an indirect approach was used. Labeling of the anti bodies with a hydrazide derivative of biotin did not affect the patter n of binding. However, significant differences were noted, when conjug ation of label was targeted to amino groups via N-hydroxy-succinimide esters of biotin and digoxigenin despite performance of the modificati on under activity-preserving conditions. Notably, the apparent strong staining of Leydig cells and nuclei of primary spermatocytes, respecti vely was not inhibitable by sugar. These differences were corroborated by a nonidentical response of the various probes in solid-phase assay s with extract proteins. Thus, care should be exercised in the interpr etation of histochemical data, obtained with this type of modified ant ibody. When these precautions are fulfilled, this immunoglobulin fract ion from human serum has the potential as an alpha-galactosyl-specific histochemical tool.