Automated muscle fiber type population analysis with ImageJ of whole rat muscles using rapid myosin heavy chain immunohistochemistry

Muscle Nerve. 2016 Aug;54(2):292-9. doi: 10.1002/mus.25033. Epub 2016 Jun 13.

Abstract

Introduction: Skeletal muscle consists of different fiber types which adapt to exercise, aging, disease, or trauma. Here we present a protocol for fast staining, automatic acquisition, and quantification of fiber populations with ImageJ.

Methods: Biceps and lumbrical muscles were harvested from Sprague-Dawley rats. Quadruple immunohistochemical staining was performed on single sections using antibodies against myosin heavy chains and secondary fluorescent antibodies. Slides were scanned automatically with a slide scanner. Manual and automatic analyses were performed and compared statistically.

Results: The protocol provided rapid and reliable staining for automated image acquisition. Analyses between manual and automatic data indicated Pearson correlation coefficients for biceps of 0.645-0.841 and 0.564-0.673 for lumbrical muscles. Relative fiber populations were accurate to a degree of ± 4%.

Conclusions: This protocol provides a reliable tool for quantification of muscle fiber populations. Using freely available software, it decreases the required time to analyze whole muscle sections. Muscle Nerve 54: 292-299, 2016.

Keywords: ImageJ; automated muscle fiber analysis; fiber population; muscle fiber type; myosin heavy chain; rat.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Diagnosis, Computer-Assisted
  • Diagnostic Imaging
  • Immunohistochemistry
  • Male
  • Muscle Fibers, Skeletal / metabolism*
  • Myosin Heavy Chains / classification
  • Myosin Heavy Chains / metabolism*
  • Rats
  • Rats, Sprague-Dawley
  • Regression Analysis

Substances

  • Myosin Heavy Chains