Skeletal muscle satellite cells are adult muscle-derived stem cells receiving 
increasing attention. Sheep satellite cells have a greater similarity to human 
satellite cells with regard to metabolism, life span, proliferation and 
differentiation, than satellite cells of the rat and mouse. We have used 2-step 
enzymatic digestion and differential adhesion methods to isolate and purify 
sheep skeletal muscle satellite cells, identified the cells and induced 
differentiation to examine their pluripotency. The most efficient method for the 
isolation of sheep skeletal muscle satellite cells was the type I collagenase 
and trypsin 2-step digestion method, with the best conditions for in vitro 
culture being in medium containing 20% FBS+10% horse serum. Immunofluorescence 
staining showed that satellite cells expressed Desmin, α-Sarcomeric Actinin, 
MyoD1, Myf5 and PAX7. After myogenic induction, multinucleated myotubes formed, 
as indicated by the expression of MyoG and fast muscle myosin. After osteogenic 
induction, cells expressed Osteocalcin, with Alizarin Red and ALP (alkaline 
phosphatase) staining results both being positive. After adipogenic induction, 
cells expressed PPARγ2 (peroxisome-proliferator-activated receptor γ2) and clear 
lipid droplets were present around the cells, with Oil Red-O staining giving a 
positive result. In summary, a successful system has been established for the 
isolation, purification and identification of sheep skeletal muscle satellite 
cells.