Bioluminescent Indicator for Highly Sensitive Analysis of Estrogenic Activity in a Cell-Based Format.
Estrogens regulate different physiological systems with wide ranges of concentrations. The rapid analysis of estrogens is crucially important for drug discovery and medical diagnosis, but quantitation of nanomolar estrogens in live cells persists as an important challenge. We herein describe a bioluminescent indicator used to detect low concentrations of estrogens quantitatively with a high signal-to-background ratio. The indicator comprises a ligand-binding domain of an estrogen receptor connected with its binding peptide, which is sandwiched between split fragments of a luciferase mutant. Results show that the indicator recovered its bioluminescence upon binding to 17β-estradiol at concentrations higher than 1.0 × 10-10M. The indicator was reactive to agonists but did not respond to antagonists. The indicator is expected to be applicable for rapid screening estrogenic compounds and inhibitors, facilitating the discovery of drug candidates in a high-throughput manner.
Department of Chemistry, School of Science, The University of Tokyo , 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
Osamu
O
Takenouchi
Department of Chemistry, School of Science, The University of Tokyo , 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
Akira
A
Kanno
Department of Chemistry, School of Science, The University of Tokyo , 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
Hideo
H
Takakura
Department of Chemistry, School of Science, The University of Tokyo , 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
Mitsuru
M
Hattori
Department of Chemistry, School of Science, The University of Tokyo , 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
Takeaki
T
Ozawa
D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D004958:Estradiol / Q000819:agonists / Q000494:pharmacology; D004967:Estrogens / Q000819:agonists / Q000494:pharmacology*; D008163:Luminescent Measurements / Q000379:methods*; D059629:Signal-To-Noise Ratio
D004967:Estrogens; D004958:Estradiol