Ab124205

The CPM cellular proteins and muscle tissues were extracted using radio-immunoprecipitation assay (RIPA) buffer and phenylmenthanesulfonyl fluoride (PMSF) protease inhibitor. The Western blot assays were carried out as previously reported^{48 (link)}. The antibodies used for Western blots were as follows: Flag tag polyclonal antibody (20543-1-AP; Proteintech, USA; 1:1000), MyHC mouse monoclonal antibody (B103; DHSB, Lowa City, IA, USA; 1:500), rabbit anti-heavy chain Myosin (ab124205; Abcam, Cambridge, UK; 1:1000), HRP-conjugated monoclonal mouse anti-glyceraldehyde-3-phosphate dehydrogenase (GAPDH) (KC-5G5; Kangchen, China; 1:10,000). HRP conjugated goat anti-rabbit IgG (A21020; Abbkine, USA; 1:10,000) and HRP conjugated goat anti-mouse IgG (A21010; Abbkine, USA; 1:10,000) were used as secondary antibodies.

Sections (14 μm) were prepared from frozen embryos, and were immunostained using the following antibodies: mouse monoclonal anti-quail cell (QCPN; Developmental Studies Hybridoma Bank; 1:100), mouse monoclonal anti-QH1 (Developmental Studies Hybridoma Bank; 1:100), mouse anti-Isl1 (Developmental Studies Hybridoma Bank; 1:100), mouse anti-MHC (MF20; Developmental Studies Hybridoma Bank; 1:100), mouse anti-sarcomeric α actinin (ab9465; Abcam; 1:100), rabbit anti-MHC (ab124205; Abcam; 1:100), rabbit anti-cardiac troponin I antibody (ab47003; Abcam; 1:100), rabbit anti-Nkx2.5 (ab35842; Abcam; 1:100). Signals were visualized with FITC (fluorescein isothiocyanate)- or TRITC-conjugated secondary antibodies specific for the appropriate species. Some sections were treated with biotin-conjugated secondary antibodies and visualized using the Cy3 streptavidin (Biolegend; 1:1000) or Dylight 488 streptavidin (Biolegend; 1:500). Nuclei were visualized with TO-PRO-3 Iodide (Molecular Probes; 1:1000). Fluorescent signals were visualized with a computer-assisted confocal microscope (Nikon ECLIPSE C2/Ti) and software (Nikon NIS-Elements AR4.100).