Ultrastructural localization of lectin receptors on the surface of the rat retinal pigment epithelium. Decreased sensitivity of the avidin-biotin method due to cell surface charge.
Monosaccharides on the apical processes of the retinal pigment epithelium were examined using lectin-affinity cytochemical methods. Lectin receptor sugars were localized with lectin-horseradish peroxidase (HRP) and lectin-ferritin conjugates as well as with biotinylated lectins, avidin, and biotinylated HRP. In contrast, only wheat germ agglutinin (WGA) receptors were identified with biotinylated WGA followed by avidin-ferritin or free avidin and biotinylated ferritin. Labeling with avidin-ferritin subsequent to biotinylated lectin treatment was dependent upon the source and lot of the reagent. These findings are similar to those reported for the endothelium of bone marrow sinusoids (Pino RM: Am J Anat, 169:259, 1984). Since both the retinal pigment epithelial and bone marrow sinusoidal surfaces are highly anionic (negative), we investigated the possibility that the charge of the lectin reagents and cell surfaces might affect the localization of monosaccharides on cell surfaces. Analytical isoelectric focusing revealed that biotinylated ferritin and some avidin-ferritins are highly anionic, while the other lectin reagents have more cationic (positive) components. Based on this information, a less charged biotinylated ferritin marker was made that made it possible to localize biotinylated lectins bound to the cell surface.
D000818:Animals; D001360:Avidin; D001710:Biotin; D007202:Indicators and Reagents; D007525:Isoelectric Focusing; D008854:Microscopy, Electron; D010857:Pigment Epithelium of Eye / Q000032:analysis / Q000648:ultrastructure*; D051381:Rats; D011919:Rats, Inbred Strains; D011920:Rats, Inbred WF; D011975:Receptors, Mitogen / Q000032:analysis*
D007202:Indicators and Reagents; D011975:Receptors, Mitogen; D001360:Avidin; D001710:Biotin