Muscle satellite cells are heterogeneous and present functional disparities, some of them behaving as multipotent stem cells. Yet their phenotype is obscure and their isolation remains elusive. The ability to purify stem cells from a wide variety of tissues by using Hoechst 33342 staining/FACS methods has permitted access to this category of cells (side population, or SP) in a manner independent of antibodies. Here, we show that the C2C12 myogenic line comprises a minor population of cells with SP phenotype. These cells are growth-arrested and delayed in their ability to differentiate. Dye efflux in C2C12-derived SPs is likely mediated by mdr1a, whose overexpression results in increased dedifferentiation. Interestingly, growth-arrested SPs rapidly appear in purified MP populations, thus suggesting a dynamic equilibrium among different states of differentiation. Finally, transcriptional profiling of C2C12-derived SP and MP cells corroborates the many similarities of SP to stem cells.