Immunofluorescence analysis of myogenic differentiation

Research output: Chapter in Book/Report/Conference proceedingChapter

Abstract

Skeletal muscle is a highly regenerative tissue that can efficiently recover from various damages caused by injuries and excessive exercises. In adult muscle, stem cells termed satellite cells are mitotically quiescent but activated upon muscle damages to enter the cell cycle as myogenic precursor cells or myoblasts. After several rounds of cell cycles, they exist the cycle and fuse to each other to form multinucleated myotubes, and eventually mature to become contractile myofibers. Satellite cells can be readily isolated from mouse skeletal muscle with enzymatic digestion and magnetic separation with antibodies against specific surface markers. C2C12 cells are an immortalized mouse myoblast cell line that is commercially available and more readily expandable than primary myoblasts. Both primary myoblasts and C2C12 cells have been extensively used as useful in vitro models for myogenic differentiation. Proper examination of this process requires monitoring specific protein expression in subcellular compartments, which can be accomplished through immunofluorescence staining. This chapter describes the workflow for the isolation of satellite cells from mouse skeletal muscle and subsequent immunofluorescence staining to assess the proliferation and differentiation of primary myoblasts and C2C12 cells.

Original languageEnglish (US)
Title of host publicationMethods in Stem Cell Biology - Part A
EditorsIlio Vitale, Gwenola Manic, Lorenzo Galluzzi
PublisherAcademic Press Inc.
Pages117-125
Number of pages9
ISBN (Print)9780323900201
DOIs
StatePublished - Jan 2022

Publication series

NameMethods in Cell Biology
Volume170
ISSN (Print)0091-679X

Bibliographical note

Funding Information:
A.A. was supported by the NIH (R01AR062142 and R21AR070319). N·K was supported by the NIH (R01GM137603 and R21AR076167). The content is solely the responsibility of the authors and does not necessarily represent the official views of the NIH.

Publisher Copyright:
© 2022 Elsevier Inc.

Keywords

  • Differentiation
  • Immunofluorescence
  • Myoblast
  • Myogenesis
  • Myotube
  • Satellite cells
  • Skeletal muscle

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