Rabbit anti α sma

For pluripotency marker stainings, iPSC colonies were passaged as described above and grown on Matrigel^TM-coated coverslips in ES medium containing, 50% MEF-conditioned media (own preparation) supplemented with 5 ng/ml FGF2 (Sigma). Colonies were then stained for alkaline phosphatase according to the manufacturer’s protocol (Millipore) or were fixed with 4% paraformaldehyde (PFA) in PBS for 10 min at RT for analysis of pluripotency markers by immunofluorescence. Fixed colonies were incubated for 2 h in blocking solution (3% normal horse serum and 0.05–0.2% Triton-X100 in PBS). Plates were incubated over night at 4 °C using the following primary antibodies: rabbit anti-Nanog (1:500), rabbit anti-Oct4 (1:1000), mouse anti-SSEA4 (1:500), mouse anti TRA-1-60 (1:500) (all from Abcam) and mouse anti-Sox2 (1:500, R&D Systems). Differentiated EBs were stained with rabbit anti-α-SMA (1:500, Sigma Aldrich), mouse anti-α-Fetoprotein (1:500, Abcam), rabbit anti-GATA4 (1:500, Abcam), and mouse anti-TUJ1 (1:1000, Covance), mouse anti-Actinin (Sigma, 1:200) and mouse anti Beta-Catenin (BD Bioscience 1:500).

HCFs were cultured on 4-well cell culture slides (30,104; SPL Life Sciences, Pocheon, Korea). After fixation in 4% paraformaldehyde, the cells were incubated with 0.1% Triton X-100 in PBS for 30 min, blocked for 1 h with 1% bovine serum albumin in PBS, and incubated with rabbit anti-KLF4 (Abcam, Cambridge, UK) and rabbit anti- α-SMA (Sigma Chemical Co, MO, USA) antibodies. The secondary antibodies were anti-rabbit IgG and anti-mouse IgG (BioLegend, CA, USA). Nuclei were counterstained with 4′,6-diamidino-2-phenylindole dihydrochloride (DAPI) (Sigma Chemical Co). Fluorescence images were obtained using an Olympus CKX41 microscope (Tokyo, Japan).