Isolation and Expansion of Muscle Satellite Cells Using the Ice-Cold Treatment Method

Satellite cells (SCs) are skeletal muscle stem cells responsible for muscle regeneration after injury. In this chapter, we describe the ice-cold treatment (ICT) method, a procedure developed for isolating and expanding highly pure populations of mouse and human SCs in vitro. The ICT method involves placing a dish containing a mixed population of adherent muscle mononuclear cells on ice for 15-30 min. This selectively causes only the SCs to detach, resulting in cell cultures with purity levels ranging from 95% to 100%. Furthermore, this method supports serial passaging, allowing prolonged expansion of SCs without negatively impacting their ability to proliferate or differentiate.