Nis elements br analysis 4.13.04 64 bit software

iPSC-CMs were cultured in RPMI with B27 supplement and 2% FBS 2 days prior to EdU staining to accelerate the proliferation. Staining was performed using the Click-it EdU Imaging Kit (Life technologies) according to the manufacturer’s instructions. EdU was incorporated for 24 h prior to experiment. Cells were fixed with 4% PFA, permeabilized with 0.5% TritonX-100 in PBS, and stained with Alexa Fluor-conjugated azide. After washing and blocking with 5% goat serum in PBST, cells were incubated with a rabbit anti-TNNT2 antibody (Abcam) and visualized with anti-rabbit Alexa Fluor-conjugated secondary antibody and Hoechst 33342. At least more than one hundred cells were counted per sample and more than three independent studies per group were performed. Measurement of the fluorescent signals was performed using NIS elements BR analysis 4.13.04 64-bit software (Nikon).