LIGHT AND ELECTRON-MICROSCOPE ANALYSIS OF LECTIN BINDING TO ADULT-RAT LIVER INSITU

Abstract

A comprehensive mapping of lectin receptors on adult rat liver in situ was performed at light and ultrastructural levels by using 12 biotin-labeled lectins and an avidin-biotin-peroxidase complex. Concanavalin A conjugated directly to peroxidase was utilized to study intracellular membrane glycoconjugates. To achieve optimal preservation of these membrane sugar moieties, several fixatives and fixation procedures were evaluated. A periodate-lysine-paraformaldehyde combination provided the best compromise between preservations of ultrastructural details and lectin-binding reactivity. Hepatocyte cell surfaces reacted intensely with concanavalin A, Lens culinaris agglutinin and Pisum sativum agglutinin (all specific for .alpha.-D-mannosyl and a-D-glucosyl groups) as well as Ricinus communis agglutinin type I (specific for .alpha. or .beta.-D-galactose) and wheat germ agglutinin (specific for neuraminic acid and .beta.-NAc-glucosaminyl groups). R. communis agglutinin and wheat germ agglutinin exhibited an extremely strong reactivity for bile canaliculi which surpassed the binding of concanavalin A, L. culinarsi agglutinin and P. sativum to these structures. Phaseolus vulgaris agglutinin (specific for .beta.-D-galactose-glucosyl-NAC and D-mannosyl groups), which exhibited a moderate binding to hepatocyte plasma membranes, reacted more strongly with the endothelium of sinusoids and portal vessels. Although all 6 of these lectins plus Bandieraea simplicifolia stained Kupffer cells, B. simplicifolia lectin (an .alpha.-D-galactosyl marker) was unique in showing a strong reactivity for only this cell type. The avidin-biotin-peroxidase procedures is a sensitive method for detection of sugar moieties on cell surfaces of rat liver at both light microscopic and EM levels. In this study, the procedure was used to localize differential binding of lectins to several anatomical structures of the organ, and furthermore, the preferential localizations of carbohydrate residues in the glycocalyx of the rat hepatocyte in situ was mapped out.