Myosin heavy chain myhc

Immunofluorescence analysis was performed on X-METs cross and longitudinal sections. X-METs were embedded in tissue-freezing medium and snap frozen in nitrogen-cooled isopentane. Samples were mounted on a cryostat and cut at 10 μm thick sections. Section was fixed with 4% PFA, washed in PBS with 1% BSA and 0.2% Triton X-100, preincubated for 1 h in 10% goat serum at RT, and incubated overnight at 4 °C with the following primary antibodies: Myosin Heavy Chain (MyHC) (Sigma-Aldrich), Connexin-43 (Cx-43) (Sigma-Aldrich) and Cardiac troponin (Troponin I) (RV-C2 Hybridoma Bank). Sections were then washed in PBS with 0.2% Triton X-100 and incubated with secondary antibody (Alexa Fluor, Life Technologies) 45 min at room temperature. Nuclei were stained using Pibenzimol bisbenzimide H33342 (HOECHST). All analyses were performed using Zeiss Confocal software (Zen 3.0 Blue edition).

Aniline, ammonium persulfate (APS), sodium
diphenylamine-4-sulfonate, hydrogen chloride (HCl), Dulbecco’s
modified Eagle’s medium (DMEM), fetal bovine serum (FBS), trypsin-ethylenediaminetetraacetic
acid (EDTA), CCK-8, Triton X-100, transforming growth factor beta
1 (TGFβ1), 4′,6-diamidino-2-phenylindole (DAPI), primary
antibodies against human alpha-smooth muscle actin (α-SMA),
and myosin heavy chain (MyHC) were obtained from Sigma-Aldrich (St.
Louis, MO, USA). Phalloidin Alexa Fluor 568, Calcein-AM, and TOTO-3
were purchased from Invitrogen (Thermo Fisher Scientific, United Kingdom).
CNA35-OG488 was obtained from the Department of Biomedical Engineering
(TU/e Eindhoven, The Netherlands), and Alexa Fluor 488 labeled goat
antimouse secondary antibody was purchased from Molecular Probes Life
Technologies.