Tissue triage and freezing for models of skeletal muscle disease

Hui Meng, Paul M.L. Janssen, Robert W. Grange, Lin Yang, Alan H. Beggs, Lindsay C. Swanson, Stacy A. Cossette, Alison Frase, Martin K. Childers, Henk Granzier, Emanuela Gussoni, Michael W. Lawlor

Research output: Contribution to journalArticle

33 Scopus citations

Abstract

Skeletal muscle is a unique tissue because of its structure and function, which requires specific protocols for tissue collection to obtain optimal results from functional, cellular, molecular, and pathological evaluations. Due to the subtlety of some pathological abnormalities seen in congenital muscle disorders and the potential for fixation to interfere with the recognition of these features, pathological evaluation of frozen muscle is preferable to fixed muscle when evaluating skeletal muscle for congenital muscle disease. Additionally, the potential to produce severe freezing artifacts in muscle requires specific precautions when freezing skeletal muscle for histological examination that are not commonly used when freezing other tissues. This manuscript describes a protocol for rapid freezing of skeletal muscle using isopentane (2-methylbutane) cooled with liquid nitrogen to preserve optimal skeletal muscle morphology. This procedure is also effective for freezing tissue intended for genetic or protein expression studies. Furthermore, we have integrated our freezing protocol into a broader procedure that also describes preferred methods for the short term triage of tissue for (1) single fiber functional studies and (2) myoblast cell culture, with a focus on the minimum effort necessary to collect tissue and transport it to specialized research or reference labs to complete these studies. Overall, this manuscript provides an outline of how fresh tissue can be effectively distributed for a variety of phenotypic studies and thereby provides standard operating procedures (SOPs) for pathological studies related to congenital muscle disease.

Original languageEnglish (US)
Article numbere51586
JournalJournal of Visualized Experiments
Issue number89
DOIs
StatePublished - Jul 15 2014

Keywords

  • Basic Protocol
  • Cell culture
  • Freezing
  • Functional testing
  • Isopentane
  • Issue 89
  • Muscle
  • Pathology
  • Tissue

ASJC Scopus subject areas

  • Neuroscience(all)
  • Chemical Engineering(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Immunology and Microbiology(all)

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  • Cite this

    Meng, H., Janssen, P. M. L., Grange, R. W., Yang, L., Beggs, A. H., Swanson, L. C., Cossette, S. A., Frase, A., Childers, M. K., Granzier, H., Gussoni, E., & Lawlor, M. W. (2014). Tissue triage and freezing for models of skeletal muscle disease. Journal of Visualized Experiments, (89), [e51586]. https://doi.org/10.3791/51586