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Journal of Histochemistry and Cytochemistry, Vol. 45, 695-702, Copyright © 1997 by The Histochemical Society, Inc.

ARTICLE

Specificity of Antibodies Against Rodent Transforming Growth Factor-{alpha} Protein

Hiroaki Aoyamaa, Hidenori Katob, and Darlene Dixona
a Laboratory of Experimental Pathology, National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina
b Department of Reproductive Physiology and Endocrinology, Medical Institute of Bioregulation, Kyushu University, Beppu, Oita, Japan

Correspondence to: Darlene Dixon, Laboratory of Experimental Pathology, MD C2-09, National Institute of Environmental Health Sciences, PO Box 12233, Research Triangle Park, NC 27709.

We found that the immunohistochemical distribution of TGF-{alpha} varied in rodent tissues depending on the antibody used, suggesting that the specificity of anti-TGF-{alpha} antibodies differs significantly. To address this issue, we compared the specificity of two representative antibodies that have been widely used to detect rodent TGF-{alpha}. In a competition study, the antibodies were preincubated with an excess of synthetic rat TGF-{alpha}34-50 and were used for staining of rat and mouse kidneys and/or uterus. The results revealed that one of the antibodies, anti-rat TGF-{alpha} polyclonal antibody, was neutralized by the peptide, whereas the other, anti-human TGF-{alpha} monoclonal antibody, was not absorbed by the peptide up to an excess of 100-fold. Western blotting analysis showed that the anti-rat TGF-{alpha} polyclonal antibody recognized both human and rat purified TGF-{alpha}. However, the anti-human TGF-{alpha} monoclonal antibody did not detect purified rat TGF-{alpha}, although the antibody reacted with mouse proteins other than TGF-{alpha} from kidneys and uterus, purified human TGF-{alpha}, and mouse carbonic anhydrase II. These data indicate that the anti-human TGF-{alpha} monoclonal antibody does not recognize rodent TGF-{alpha} under our experimental conditions and suggest that distribution of TGF-{alpha} in rodent tissues may need to be reexamined. (J Histochem Cytochem 45:695-701, 1997)

Key Words: TGF-{alpha}, anti-TGF-{alpha} antibody, immunohistochemistry, competition study, Western blotting, rats and mice


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