Isolation and grafting of single muscle fibres

Satellite cells are mononucleate muscle precursor cells resident beneath the basal lamina, which surrounds each skeletal muscle fibre. Normally quiescent in adult muscle, in response to muscle damage satellite cells are activated and proliferate to generate a pool of muscle precursor cells, which subsequently differentiate and fuse together to repair and replace terminally differentiated muscle fibre syncytia. Cells prepared by enzymatic digestion of whole muscle tissue are likely to contain myogenic cells derived both from the satellite cell niche and from other populations in the muscle interstitium and vasculature. Single muscle fibre preparations, in which satellite cells retain their normal anatomical position beneath the basal lamina, are free of interstitial and vascular tissue and can therefore be used to investigate satellite cell behaviour in the absence of other myogenic cell types. Here, we describe methods for the isolation of viable muscle fibres and for grafting of muscle fibres and their associated satellite cells into mouse muscles to assess the contribution of satellite cells to muscle regeneration.