DISTRIBUTION OF CARBOHYDRATES IN THE ZONA-PELLUCIDA OF HUMAN OOCYTES

The purpose of the present study was to investigate the distribution p attern of carbohydrates in the zona pellucida of human oocytes using l ectins and ruthenium red as histochemical probes. For lectin analyses, oocytes that failed to undergo fertilization following in vitro insem ination were collected washed, fixed with glutaraldehyde and embedded in araldite. For ruthenium red labelling, the oocytes were fixed with glutaraldehyde containing ruthenium red, post-fixed with OsO4 and embe dded in araldite. Araldite sections (1 mu m) were de-resined with sodi um methoxide, rehydrated, labelled with ten different biotinylated lec tins as probes and avidin-biotin-peroxidase complex as visualant, and examined under a light microscrope. The zonae pellucidae of all oocyte s studied exhibited a common lectin-binding pattern, expressed in inte nse binding of lectins from Concanavalia ensiformis (ConA), Lens culin aris (LCA), Ricinus communis (RCA-I), wheat germ agglutinin (WGA), and of succinylated WGA (S-WGA). Peanut lectin (PNA) bound to the zona pe llucida only after neuraminidase treatment, whereas the lectins from G riffonia simplisifolia (GS-I), Dolichos biflorus (DBA), Ulex europhaeu s (UEA-I) and soybean (SBA) did not bind at all. There was almost no b inding of ruthenium red to the matrix of the zona pellucida. The resul ts indicate that the human zona pellucida is characterized by normally exposed mannosyl, N-acetylglucosaminyl and beta-galactosyl residues. In addition, it contains masked beta Gal-(1-3)GalNAc sugar sequences t hat can be exposed only after removing terminal sialic acid residues. The presence of sialic acid in the human zona pellucida, which is not expressed as an increase in the binding capacity of the polycationic p robe (ruthenium red), has not been reported in any of the mammalian zo nae pellucidae previously studied.