Isolation and Sorting of Murine Satellite Cells

Satellite cells (SCs) were sorted as previously reported (Ryall et al., 2015 (link)). Briefly, hindlimb muscles from 6-week-old male mice were harvested, and blood, tendons, lipid tissues and nerves were removed. The muscles were finely chopped until a slurry was generated. The slurry was digested with 0.2% collagenase II and 0.1% dispase at 37°C for 60 min. The slurry was centrifuged, filtered and incubated with a primary antibody cocktail mixture as follows: APC-labeled α7-integrin, PE-labeled CD11b, CD31, CD45, and Sca1 and 7-AAD. Sorting was conducted on SH800S (SONY) and the gating strategy was based on positive α7-integrin, negative 7-AAD and CD11b/CD31/CD45/Sca1 staining.