Protein and mRNA levels of ligand bound ERα in MCF-7 cells. A) Chemical structure of 17β-estradiol, SERMs (OHT, RU39) and SERDs (ICI, RU58). B) Western blot and quantification of ERα protein levels after 1 h, 4 h and 16 h treatment with 10 nM E2, 1 μM ICI, 1 μM RU58, 1 μM OHT and 1 μM RU39 relative to ERα protein levels in untreated (EtOH) MCF-7 cells. 2 μg of total protein were loaded. Relative protein levels from three independent experiments were quantified. C) Total RNA was extracted from MCF-7 cells treated or not for 16 h with 10 nM E2 or anti-estrogens. Relative expression level of the ESR1 and GREB1 genes was analyzed by qRT-PCR. TBP served as an internal control (see Methods). Data shown are an average of three independent experiments, error bars represent ± S.E. mean.