title: Localization of a wide-ranging panel of antigens in the rat retina by immunohistochemistry: comparison of Davidson's solution and formalin as fixatives.
abstract: The preferred fixative for whole eyes is Davidson's solution, which provides optimal tissue preservation while avoiding retinal detachment. Hitherto, the compatibility of Davidson's solution with immunohistochemistry has been largely untested. The goal of the present study was to compare the immunolabeling patterns of a wide-ranging panel of commercially available, previously validated antibodies in formalin- and Davidson's-fixed retinas. Immunohistochemistry was performed in normal pigmented rat eyes and, to facilitate localization of inducible proteins, eyes injected with the bacterial toxin lipopolysaccharide or subjected to laser-induced photoreceptor damage. Specificity of labeling was judged by the morphology and distribution of immunopositive cells, by the absence of signal in appropriate controls, and by comparison with expected staining patterns. Retinas fixed in formalin displayed only adequate morphological integrity but were highly compatible with all 39 antibodies evaluated. Retinas fixed in Davidson's solution displayed morphological integrity superior to those fixed in formalin. Generally, the cellular and subcellular patterns and intensities of immunoreactivities obtained with each fixative were identical; however, Davidson's fixative was less compatible with certain antibodies, such as the neurotransmitter γ-aminobutyric acid, the microglial marker iba1, the macroglial stress protein nestin, and the small heat shock proteins Hsp27 and αB-crystallin, shortfalls that somewhat temper enthusiasm concerning its use.
authors.affiliation: Ophthalmic Research Laboratories, South Australian Institute of Ophthalmology, Hanson Institute Centre for Neurological Diseases, Adelaide, Australia. glyn.chidlow@health.sa.gov.au
authors.forename: Glyn
authors.initials: G
authors.lastname: Chidlow
authors.forename: Mark
authors.initials: M
authors.lastname: Daymon
authors.forename: John P M
authors.initials: JP
authors.lastname: Wood
authors.forename: Robert J
authors.initials: RJ
authors.lastname: Casson
mesh_terms: D000818:Animals; D000906:Antibodies / Q000276:immunology; D000941:Antigens / Q000032:analysis* / Q000276:immunology; D002135:Calcium-Binding Proteins / Q000032:analysis / Q000276:immunology; D003459:Crystallins / Q000032:analysis / Q000276:immunology; D005404:Fixatives / Q000737:chemistry*; D005557:Formaldehyde / Q000737:chemistry*; D055551:HSP27 Heat-Shock Proteins / Q000032:analysis / Q000276:immunology; D007150:Immunohistochemistry / Q000379:methods*; D007381:Intermediate Filament Proteins / Q000032:analysis / Q000276:immunology; D008070:Lipopolysaccharides / Q000276:immunology; D008840:Microfilament Proteins / Q000032:analysis / Q000276:immunology; D008869:Microtubule-Associated Proteins / Q000032:analysis / Q000276:immunology; D009419:Nerve Tissue Proteins / Q000032:analysis / Q000276:immunology; D064231:Nestin; D051381:Rats; D012160:Retina / Q000276:immunology*; D016707:Tissue Fixation / Q000379:methods; D005680:gamma-Aminobutyric Acid / Q000032:analysis / Q000276:immunology
chemical_list: C100745:Aif1 protein, rat; D000906:Antibodies; D000941:Antigens; D002135:Calcium-Binding Proteins; D003459:Crystallins; D005404:Fixatives; D055551:HSP27 Heat-Shock Proteins; D007381:Intermediate Filament Proteins; D008070:Lipopolysaccharides; D008840:Microfilament Proteins; D008869:Microtubule-Associated Proteins; D009419:Nerve Tissue Proteins; C577486:Nes protein, rat; D064231:Nestin; C530883:cryaB protein, rat; D005557:Formaldehyde; D005680:gamma-Aminobutyric Acid