Development of Simple, High-sensitivity, Precise Assay for Estrogenic Activity Using New Screening Mammalian Cell (CHOOSER)

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  • 新しいアッセイ用ほ乳類細胞（ＣＨＯＯＳＥＲ）を用いた簡便，高感度で再現性の良いエストロゲン様活性測定方法の開発
  • 新しいアッセイ用哺乳類細胞(CHOOSER)を用いた簡便,高感度で再現性の良いエストロゲン様活性測定方法の開発
  • アタラシイ アッセイヨウホニュウルイ サイボウ CHOOSER オ モチイタ カンベン コウカンド デ サイゲンセイ ノ ヨイ エストロゲン ヨウ カッセイ ソクテイ ホウホウ ノ カイハツ

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Abstract

Several in vitro assays have been developed to assess the activity of estrogenic substances. Recently, a yeast two-hybrid screen or a MCF-7 (human breast cancer cell) screen has been adopted mainly for screening chemicals and environmental estrogenic activity. However a yeast two-hybrid screen does not assay using an animal cell. MCF-7 screen is a complicated assay and the cell used proliferates depending on estrogen, thus obstructing a precise assay for estrogenic activity. In addition both screens need long incubation period (4-6 days) for a highly sensitive assay. The CHOOSER is a Chinese hamster ovary (CHO) cell transformed with the gene encoding the human estrogen receptor (ER α) and an estrogen responsive promoter linked to a reporter gene which was developed by Suzuki et. al. at Osaka prefectural institute of public health. When the reporter gene is activated by estrogenic substances, is expressed, producing alkaline phosphatase which is secreted into the medium. The CHOOSER can assay high sensitively (under 10-11M as 17 β-estradiol) at any time in a short incubation period (2 days). The number of the CHOOSER can be counted precisely because a CHO is dissociated the aggregates easily with trypsin and form a single cell suspension. And the CHOOSER can proliferate in the medium without estrogen, therefore it is possible to assay for estrogenic activity more proliferate in the medium without estrogen, therefore it is possible to assay for estrogenic activity more precisely. The CHOOSER assay will be a useful prescreening method for estrogenic activity before in vivo assay.

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