Cardiomyocyte Differentiation from Human iPSCs

Human iPSCs at 60 to 80% confluency were differentiated into cardiomyocytes as previously described (61 (link)). Briefly, cells were cultured in CDM3 media supplemented with 4 to 6 μM CHIR99021 (Selleckchem) for 48 hours (days 1 to 2) and then CDM3 media supplemented with 2 μM WNT-C59 (Selleckchem) for 48 hours (days 3 to 4). Starting on day 5, cells were cultured in basal media [RPMI 1640 (Gibco) supplemented with B-27 supplement (Thermo Fisher Scientific)] for 6 days (days 5 to 10). On day 10, cells were cultured in selective media [RPMI 1640, without glucose (Gibco), supplemented with B-27 supplement] for 10 days (days 11 to 20) and then basal media thereafter. Cardiomyocytes were used for experiments on day 30 and harvested using TrypLE Express (Thermo Fisher Scientific).