Las af version lite 2

Myogenic purity and myoblasts morphology were observed by the immunocytochemistry method using a mouse monoclonal anti-Desmin antibody (D33; Dako, Produktionsvej, Denmark). Myoblasts were plated in μ-Slide 8 well (ibidi, Martinsried, Germany) at a density of 1×10⁴ cells per well. The cells were fixed in cold ethanol. Then, anti-Desmin antibody (1:50) and Alexa Fluor 488 goat anti-mouse (Life Technologies, Carlsbad, CA, USA) were used to incubate the myoblasts in sequence. Nuclei were visualized using Hoechst 33342 (Life Technologies, Carlsbad, USA). The slides were then viewed under a Confocal Laser Scanning Microscope Leica TCS SP5 II, and data were acquired using LAS AF version Lite 2.6 software (Leica Microsystems, Wetzlar, Germany). To determine the percentage of desmin-positive cells, a minimum of 50 cells were counted in three independent cultures. In addition, the morphological changes of myoblasts were observed, while the width and length of myoblasts were visualized and measured using LAS AF version Lite 2.6 software (Leica Microsystems, Wetzlar, Germany). For each group of cells, at least 30 cells were analyzed.