Detection of binding sites for biotinylated neoglycoproteins and heparin (endogenous lectins) during cerebellar ontogenesis in the rat

Endogenous carbohydrate-binding sites were studied during rat cerebellar development on sections of fixed tissue using synthetic tools, biotinylated neoglycoproteins, in conjunction with subsequent avidinperoxidase staining. Neoglycoproteins were constructed by chemically coupling the histochemically pivotal carbohydrate moieties to an inert carrier protein. The sugar part of the neoglycoproteins included common constituents of the carbohydrate part of cellular glycoconjugates, namely mannose, galactose, fucose, N-acetyl-glucosamine, N-acetylgalactosamine and N-acetyl-neuraminic acid to probe for the presence of respective endogenous receptors. Heparin was biotinylated after mild cyanogen bromide activation and aminoalkylation. Specific positive reactions were obtained for all neoglycoproteins and heparin. The staining pattern with the individual probes disclosed variable developmental regulation. Consequently, these results suggest that recognition processes during cerebellar development may include several types of carbohydrate determinants. In two instances, the binding of neoglycoproteins could be compared to endogenous lectin-specific antibodies. Despite a significant extent of accordance the comparison revealed notable differences. These differences were attributed primarily to fixation and the presence of physiological ligands that can mask the active endogenous carbohydrate-binding proteins. In any case, histochemical application of labeled neoglycoproteins is valuable to discern the presence, localization and developmental pattern of binding sites for the carbohydrate part of glycoconjugates, on which further biochemical and cell biological studies can consequently be based.