Post-embedding localization of glycoconjugates by means of lectins on thin sections of tissues embedded in LR white.
A simple post-embedding technique for the electron microscopical detection of lectin-binding sites using thin sections of tissues embedded in the resin LR White is described. With this technique, no prior etching of the sections is necessary. The cellular fine structure is well preserved and permits close correlation of the labelling to distinct cellular compartments. After mild aldehyde fixation (4% formaldehyde and 0.5% glutaraldehyde for 30 min), enterocyte brush border, vesicles and lysosomes as well as goblet cell Golgi apparatus and mucin are intensely stained after 30-60 min. The hydrophilia and penetrability of LR White is shown by the formation of oxidized diaminobenzidine reaction product arising from horseradish peroxidase-conjugated lectins. The precipitate not only covers the surface of the sections but is also formed within the resin, as is revealed on cross-sections through thin and semithin sections. The addition of 0.2 M solutions of the appropriate inhibitory sugars prevented staining, which indicates a specific binding. Examples are given of the binding of gold-, ferritin- and peroxidase-conjugated lectins for the purpose of detecting glycoconjugates in various intracellular compartments.
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Ellinger
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Pavelka
D015100:3,3'-Diaminobenzidine; D000180:Acrylic Resins; D000818:Animals; D004386:Duodenum / Q000032:analysis / Q000648:ultrastructure; D005260:Female; D005293:Ferritins; D006046:Gold; D006056:Golgi Apparatus / Q000032:analysis; D006651:Histocytochemistry / Q000379:methods; D006735:Horseradish Peroxidase; D007082:Ileum / Q000032:analysis / Q000648:ultrastructure; D008871:Microvilli / Q000032:analysis; D009077:Mucins / Q000032:analysis; D051381:Rats; D011919:Rats, Inbred Strains; D011975:Receptors, Mitogen / Q000032:analysis*; D013194:Staining and Labeling; D014132:Trachea / Q000032:analysis / Q000648:ultrastructure
D000180:Acrylic Resins; D009077:Mucins; D011975:Receptors, Mitogen; D015100:3,3'-Diaminobenzidine; D006046:Gold; D005293:Ferritins; C048707:LR white; D006735:Horseradish Peroxidase