Development of a Competitive Enzyme Immunoassay for Detection of Capacity of Chemicals to Bind Quail Estrogen Receptor α and β

Shinobu Maekawa, Makoto Nishizuka, Shiro Heitaku, Masaaki Kunimoto, Jun-ichi Nishikawa, Kouhei Ichikawa, Kiyoshi Shimada, Masayoshi Imagawa
2004 Journal of health science  
In vitro binding assays are useful in the initial screening of endocrine disrupting chemicals. Such assays should be applied to the estrogen receptors (ER) of not only humans but also wildlife. As a system for birds is yet to established, we expressed the ligand binding domain (LBD) of quail ERα and ERβ as a fusion protein with glutathione S-transferase and using these proteins, developed two systems (a competitive enzyme immunoassay and a fluorescence polarization assay) for assaying the
more » ... assaying the capacity to bind ERs in vitro. Moreover, 20 test chemicals selected by Ministry of the Environment of Japan were evaluated in terms of binding ability. Both systems worked well, the competitive enzyme immunoassay proving especially powerful, since it needs no special equipment. This system is applicable to other species including fish, amphibians and reptiles when information on the LBD of ER is available. Key words ---endocrine disrupting chemicals, estrogen receptor, enzyme immunoassay, quail, endocrine disruptor, in vitro binding assay completed. 5) For the initial screening in vitro, the receptor binding assay is often utilized. The yeast two-hybrid assay is particularly useful, since it is easy and relatively cheap to perform, and also no special equipment is needed. 6,7) This method is based on the interaction between the ligand binding domain (LBD) in the hormone receptor and the coactivator in the ligand-dependent manner. 6) Recently, it was proposed that the effect of EDCs should be considered not only in humans but also in wildlife, and indeed adverse effects on humans and wildlife were reported. However, information on coactivators is restricted to specific species, such as humans, rats and mice. 1,2) Therefore, the yeast two-hybrid assay does not seem to be a suitable method for use in vitro among various species.
doi:10.1248/jhs.50.25 fatcat:qc5kbczni5aqrewitwzarqpkpa