Un scan it graph digitizer software 7

Whole cell extracts (WCE) were prepared in RIPA buffer (Sigma) with added phosphatase and complete protease inhibitors (Roche). Protein concentration was determined using the Bio-Rad DC protein assay (Bio-Rad). Where indicated, 10 or 15 μg of protein was boiled for 5 min, electrophoresed on 10% SDS-PAGE gels and electroblotted on PVDF membranes (Bio-Rad) for western blotting with the following antibodies: ERα (G-20, Santa Cruz Biotechnology, Santa Cruz, CA, USA), DICER1, PTEN (Cell Signaling, Danvers, MA, USA) and β-actin (loading control; Sigma). Bands were visualized using Carestream Image Station 4000R PRO with Carestream Molecular Imaging Software Version 5.0.2.30 (Carestream Health, Inc., New Haven, CT, USA) and quantified by UN-SCAN-IT Graph Digitizer Software 7.1 (Silk Scientific, Orem, UT, USA). The values were normalized to loading control and the normalized values for vehicle-treated and/or control-transfected cells were set to 1 for comparison within each cell line.