Immunostaining and Muscle Fiber Size Analysis

The mouse monoclonal antibodies specific for eMyHC (BF45) and type 1 MyHC (BAF8) were purchased from Developmental Studies Hybridoma Bank. Tissues were cryo-sectioned at 10 μm and immunostained using antibodies specific for eMyHC (1:10) or type 1 MyHC (1:5) overnight at 4 °C and processed as before^{69 (link)}. For fiber size measurements, the tissue sections were stained with WGA, (fluorophore conjugated, 1:100, Cat.# W11262, Life Technologies). Images were obtained using a Zeiss LSM 510 on Zeiss Axiovert 100 M Base and processed using NIS Elements. The minimal “Feret’s” diameter variance coefficients of the muscle fiber size was calculated on the WGA stained sections using the ImageJ 1.43u program.