Development of Standardized in Vitro Assay System for Estrogen Receptors and Species Specificity of Binding Ability of 4-Nonylphenol and p-Octylphenol

Nishizuka Makoto, Heitaku Shiro, Maekawa Shinobu, Nishikawa Jun-ichi, Imagawa Masayoshi

doi:10.1248/jhs.50.511

The in vitro binding assay seems to be a useful first screening method for endocrine disrupting chemicals. The various methods have been developed and applied to the testing of chemicals. Although these assays should be applied to estrogen receptors (ER) of not only humans but also wildlife, a standardized system is yet to be established. Furthermore, a method for Xenopus ER is not yet developed. We previously expressed the ligand-binding domain (LBD) of quail ERα and ERβ as a fusion protein with glutathione S-transferase, and developed a competitive enzyme immunoassay for detecting the capacity of chemicals to bind ERs. It seems that this system is a powerful tool, since it needs no special equipment. In this report, we first produced ER-LBD protein of human, Xenopus and medaka as well as quail. Then, we established a competitive enzyme immunoassay for these ERs as a standardized method, and compared the species specificity of the ability of 4-nonylphenol and p-octylphenol to bind ERs. Although a significant difference was not detected among ERβ of human, quail and medaka, 4-nonylphenol and p-octylphenol exhibited the higher affinity for the medaka ERα than human ERα. These results indicate the species specificity of the capacity of chemicals to bind ERs. <br>

Development of Standardized in Vitro Assay System for Estrogen Receptors and Species Specificity of Binding Ability of 4-Nonylphenol and p-Octylphenol

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