Post-embedding tem signal-to-noise ratio of S-100.
We assessed the reactivity of purified S-100 antiserum in immuno-electron microscopy by counting the number of gold particles per microns 2 over inner ear tissues embedded in different media. Sections containing predominantly Schwann's cell cytoplasm and nucleus, afferent fiber axoplasm and myelin sheath of chick cochleae were reacted with anti-S-100 IgG, an antibody to a calcium binding protein of neuronal tissues, then labeled with anti-IgG-gold conjugate. This investigation was conducted because previously published procedures, unmodified, did not yield acceptable results. Preparation of all specimens was identical. Only the medium (PolyBed 812, Araldite or Spurr epoxies; and LR White, LR Gold or Lowicryl plastics) was changed. The medium was made the changing variable because antigens available in post-embedding immuno-electron microscopy are decreased by heat, either used and/or released during polymerization of the embedding medium. The results indicate that: (a) none of the embedding media above provided optimal signal-to-noise ratio for all parts of the nerve stained in the same section; (b) aggregation of gold particles over cells was highest in embedding media with high background labeling over areas devoid of tissue (noise); (c) aggregation occurred randomly throughout both cellular and acellular regions; and (d) particles aggregated less and were distributed more evenly in tissues from media yielding good ultrastructural integrity.
Fermin
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Tulane University School of Medicine, Department of Pathology, New Orleans, LA 70112-2699.
Lee
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DH
Martin
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D000818:Animals; D002467:Cell Nucleus / Q000737:chemistry; D002645:Chickens; D003051:Cochlea / Q000294:innervation* / Q000648:ultrastructure; D003056:Cochlear Nerve / Q000737:chemistry* / Q000648:ultrastructure; D003593:Cytoplasm / Q000737:chemistry; D007106:Immune Sera; D007074:Immunoglobulin G; D007150:Immunohistochemistry; D008689:Methacrylates / Q000737:chemistry; D016253:Microscopy, Immunoelectron; D009186:Myelin Sheath / Q000737:chemistry*; D009475:Neurons, Afferent / Q000737:chemistry; D016613:Plastic Embedding; D009418:S100 Proteins / Q000032:analysis* / Q000276:immunology; D012583:Schwann Cells / Q000737:chemistry*; D016610:Tissue Embedding
D007106:Immune Sera; D007074:Immunoglobulin G; D008689:Methacrylates; D009418:S100 Proteins