title: Isolation, Culture, Functional Assays, and Immunofluorescence of Myofiber-Associated Satellite Cells.
abstract: Adult skeletal muscle stem cells, termed satellite cells, regenerate and repair the functional contractile cells in adult skeletal muscle called myofibers. Satellite cells reside in a niche between the basal lamina and sarcolemma of myofibers. Isolating single myofibers and their associated satellite cells provides a culture system that partially mimics the in vivo environment. We describe methods for isolating and culturing intact individual myofibers and their associated satellite cells from the mouse extensor digitorum longus muscle. Following dissection and isolation of individual myofibers we provide protocols for myofiber transplantation, satellite cell transfection, immune detection of satellite cell antigens, and assays to examine satellite cell self-renewal and proliferation.
authors: Department of Molecular, Cellular and Developmental Biology, University of Colorado at Boulder, Boulder, CO, 80309, USA.
authors: Thomas O
authors: TO
authors: Vogler
authors: Department of Molecular, Cellular and Developmental Biology, University of Colorado at Boulder, Boulder, CO, 80309, USA.
authors: Katherine E
authors: KE
authors: Gadek
authors: Department of Molecular, Cellular and Developmental Biology, University of Colorado at Boulder, Boulder, CO, 80309, USA.
authors: Adam B
authors: AB
authors: Cadwallader
authors: Department of Molecular, Cellular and Developmental Biology, University of Colorado at Boulder, Boulder, CO, 80309, USA.
authors: Tiffany L
authors: TL
authors: Elston
authors: Department of Molecular, Cellular and Developmental Biology, University of Colorado at Boulder, Boulder, CO, 80309, USA. Olwin@colorado.edu.
authors: Bradley B
authors: BB
authors: Olwin
mesh_terms: D000818:Animals; D018929:Cell Culture Techniques; D002454:Cell Differentiation; D049109:Cell Proliferation; D002469:Cell Separation / Q000379:methods*; D005455:Fluorescent Antibody Technique; D051379:Mice; D018482:Muscle, Skeletal / Q000166:cytology*; D032496:Satellite Cells, Skeletal Muscle / Q000166:cytology* / Q000378:metabolism* / Q000637:transplantation; D014162:Transfection
keywords: AraC; CFDA-SE; EdU; Extensor digitorum longus; Immunostaining; Mouse; Muscle stem cells; Myofiber isolation; Proliferation; Satellite cells; Self-renewal; Skeletal muscle; Tibialis anterior; Transfection; Transplant