Using a Whole-mount Immunohistochemical Method to Study the Innervation of the Biliary Tract in Suncus murinus

J Vis Exp. 2017 Jun 15:(124):55483. doi: 10.3791/55483.

Abstract

This work describes the whole-mount immunohistochemistry staining method in detail, using neurofilament protein antibody to label the innervation of the biliary tract in Suncus murinus (S. murinus ). First, the specimen was dissected from S. murinus and fixed in 4% paraformaldehyde (PFA). Second, an enzymatic treatment and potential endogenous peroxidase inactivation were performed. The specimen was then exposed to the primary antibody, anti-neurofilament protein antibody, for 3-6 days. It was then incubated with the secondary antibody conjugated with horseradish peroxidase. The color reaction was revealed by reacting the specimen with a 3,3'-diaminobenzidine (DAB) substrate. This method can be applied to analyze the innervation of all visceral organs. Furthermore, this protocol can also be adapted to test other neuronal antibodies, but optimization of the antibodies should be done first. This method was originally introduced by Kuratani and Tanaka1,2,3.

Publication types

  • Video-Audio Media

MeSH terms

  • Animals
  • Antibodies, Monoclonal / chemistry
  • Biliary Tract / anatomy & histology
  • Biliary Tract / innervation*
  • Female
  • Horseradish Peroxidase / chemistry
  • Immunohistochemistry / methods*
  • Male
  • Neurofilament Proteins / metabolism
  • Neurons / metabolism
  • Peripheral Nerves / metabolism
  • Shrews* / anatomy & histology
  • Staining and Labeling

Substances

  • Antibodies, Monoclonal
  • Neurofilament Proteins
  • Horseradish Peroxidase