Protocol for the isolation of mouse muscle stem cells using fluorescence-activated cell sorting.

Muscle stem cells (MuSCs) are the building blocks for regenerating skeletal muscle after trauma. If we intend to maximize the therapeutic potential of MuSCs, we must further study their molecular and functional properties. Here, we present a protocol for the isolation of mouse MuSCs via a two-step enzymatic and mechanical dissociation of skeletal muscle coupled with fluorescence-activated cell sorting (FACS). FACS-isolated MuSCs can be used for various downstream applications including cell culture, cell transduction, immunofluorescence, and gene expression assays. For complete details on the use and execution of this protocol, please refer to Almada et al. (2021). H1: Optimized technique to isolate mouse MuSCs by FACS H2: Ex vivo culturing approaches to study MuSC fate decisions H3: Immunofluorescence and RNA analysis of MuSCs H4: Approaches for transduction and transplantation of MuSCs Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. Muscle stem cells (MuSCs) are the building blocks for regenerating skeletal muscle after trauma. If we intend to maximize the therapeutic potential of MuSCs, we must further study their molecular and functional properties. Here, we present a protocol for the isolation of mouse MuSCs via a two-step enzymatic and mechanical dissociation of skeletal muscle coupled with fluorescence-activated cell sorting (FACS). FACS-isolated MuSCs can be used for various downstream applications including cell culture, cell transduction, immunofluorescence, and gene expression assays.

肌肉干细胞（MuSCs）是创伤后骨骼肌再生的基石。如果我们想要最大限度地发挥肌肉干细胞的治疗潜力，就必须进一步研究它们的分子和功能特性。在此，我们介绍一种通过骨骼肌的两步酶解和机械解离并结合荧光激活细胞分选（FACS）来分离小鼠肌肉干细胞的实验方案。通过FACS分离的肌肉干细胞可用于多种下游应用，包括细胞培养、细胞转导、免疫荧光和基因表达分析。 如需该实验方案使用和实施的完整细节，请参考阿尔马达等人（2021年）的研究。 H1：通过FACS分离小鼠肌肉干细胞的优化技术 H2：用于研究肌肉干细胞命运决定的体外培养方法 H3：肌肉干细胞的免疫荧光和RNA分析 H4：肌肉干细胞的转导和移植方法 出版者注：进行任何实验方案都需要遵守当地机构关于实验室安全和伦理的指导方针。 肌肉干细胞（MuSCs）是创伤后骨骼肌再生的基石。如果我们想要最大限度地发挥肌肉干细胞的治疗潜力，就必须进一步研究它们的分子和功能特性。在此，我们介绍一种通过骨骼肌的两步酶解和机械解离并结合荧光激活细胞分选（FACS）来分离小鼠肌肉干细胞的实验方案。通过FACS分离的肌肉干细胞可用于多种下游应用，包括细胞培养、细胞转导、免疫荧光和基因表达分析。