Detection of estrogen receptor (ER) in the rat brain using rat anti-ER monoclonal IgG with the unlabeled antibody method.
Application of Sternberger's unlabeled antibody enzyme method for detection of the estrogen receptor (ER) using a rat primary antibody with rat tissues has been discouraged, presumably because nonspecific staining of endogenous IgG was expected with the required anti-rat IgG bridging antibody. Because the blood-brain barrier greatly reduces immunoglobulin infiltration into the brain, we hypothesized that rat brain tissue could be specifically immunostained using rat IgG primary antibodies. A rat monoclonal anti-ER antibody (H222) specifically stained ERs in the brains of ovariectomized but not in ovariectomized estrogen-treated rats. In contrast, the uterus, a well-perfused target organ stained intensely in a nonspecific fashion. Dense populations of estrogen receptors were observed in the medial preoptic area, the bed nucleus of the stria terminalis, and the arcuate and ventromedial nuclei of the hypothalamus. A monoclonal rat IgG directed against alpha-tubulin labeled primarily cortical dendrites quite distinct from the neuronal nuclei that are the primary antigenic sites for the estrogen receptor antibody. These results confirm that the sensitive unlabeled antibody method can be applied to rat brain tissues, even when the primary antibody is rat IgG and that labeling of endogenous IgG may be used as a simple method to evaluate the integrity of the blood brain barrier.
Department of Health and Human Services, Food and Drug Administration, Jefferson, AR 72079.
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Henry
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Medlock
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DM
Sheehan
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Scallet
D000818:Animals; D000911:Antibodies, Monoclonal; D001921:Brain / Q000378:metabolism*; D001923:Brain Chemistry; D005260:Female; D007031:Hypothalamus / Q000378:metabolism; D007150:Immunohistochemistry / Q000379:methods*; D051381:Rats; D011919:Rats, Inbred Strains; D011960:Receptors, Estrogen / Q000032:analysis* / Q000276:immunology; D014404:Tubulin / Q000032:analysis / Q000276:immunology; D014599:Uterus / Q000378:metabolism
D000911:Antibodies, Monoclonal; D011960:Receptors, Estrogen; D014404:Tubulin