A rationale for SDS-PAGE of MHC isoforms as a gold standard for determining contractile phenotype.

: We believe that a better “gold standard” withregard to assessing fiber type, specifically the contractile phe-notype, is with SDS-PAGE separation of MHC isoforms per-formed, optimally, on single isolated muscle fibers. The met-abolic and contractile phenotype must be assessed separately,and the electrophoretic approach offers distinct advantages toATPase and immunohistochemistry (IHC). While the lattermethods are probably the most commonly used, there arelimitations to these approaches described by Booth et al. (1).Many muscle fibers express heterogeneity of MHC isoforms.ATPase and IHC provide information only on the presence ofa given isoform, not its proportional contribution. These meth-ods are therefore only informative from a qualitative perspec-tive.Our suggested methodology is feasible for most labs toperform and, importantly, is quantitative. It allows fordetermination of precise proportions of all MHC isoforms ina given muscle fiber. Additionally, it provides a definitivemeans for identifying the IIx MHC isoform, something thatis currently problematic with respect to ATPase and IHCtechniques. Neonatal and embryonic MHCs can also bedistinctly separated and quantified by electrophoresis, whenperformed under optimized conditions (6). Another specificadvantage of this methodology is that it allows for theprediction of functional properties of the muscle, i.e., force-velocity relationship (4, 5). SDS-PAGE has been defini-tively shown to be a method sensitive to subtle or markeddifferences in MHC isoform expression in response toaltered loading and hormone states, during development,and between muscles of different fiber types (2–6). Toprove that interventions impact on contractile phenotype,MHC isoform composition should be determined by theelectrophoretic approach.