title: A simple heterogeneous one-step assay for screening estrogenic compounds.
abstract: Estrogen receptor (ER) modulators are a serious health issue but estrogenic compounds, especially antagonists of ER function, are widely screened for in search of novel therapeutics against hormonal diseases such as the breast cancer. Here we report a novel and a simple bioassay for estrogenic and anti-estrogenic compounds based on ligand-dependent recruitment of ER co-activator steroid receptor co-activator 1 (SRC-1) to purified Renilla luciferase-tagged ERα. In this assay, in vivo-biotinylated (E. coli) SRC-1, purified Renilla luciferase-ERα, and the analyte sample are mixed and incubated for 2 h in a streptavidin-coated microtiter wells, and after one washing step, luminescence is measured with a simple instrument. The assay does not require chemical labeling of the components and shows good sensitivity (25 pM E(2)) and wide dynamic range of more than four orders of magnitude.
authors.affiliation: Department of Biotechnology, University of Turku, Turku, Finland. tuomas.huovinen@utu.fi
authors.forename: Tuomas
authors.initials: T
authors.lastname: Huovinen
authors.forename: Kalle
authors.initials: K
authors.lastname: Rytkönen
authors.forename: Urpo
authors.initials: U
authors.lastname: Lamminmäki
authors.forename: Teijo
authors.initials: T
authors.lastname: Pellinen
mesh_terms: D001681:Biological Assay / Q000379:methods*; D004305:Dose-Response Relationship, Drug; D052244:Endocrine Disruptors / Q000032:analysis* / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D004958:Estradiol / Q000032:analysis / Q000378:metabolism; D004965:Estrogen Antagonists / Q000032:analysis* / Q000378:metabolism; D047628:Estrogen Receptor alpha / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004967:Estrogens / Q000032:analysis* / Q000378:metabolism; D006801:Humans; D049410:Luciferases, Renilla / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D056920:Nuclear Receptor Coactivator 1 / Q000737:chemistry / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012680:Sensitivity and Specificity
chemical_list: D052244:Endocrine Disruptors; D004965:Estrogen Antagonists; D047628:Estrogen Receptor alpha; D004967:Estrogens; D011993:Recombinant Fusion Proteins; D004958:Estradiol; D049410:Luciferases, Renilla; C538859:NCOA1 protein, human; D056920:Nuclear Receptor Coactivator 1