To understand the biological relationships among various molecules, it is necessary to define the cellular expression patterns of multiple genes and gene products. Relatively simple methods for performing multi-label immunohistochemical detection are available. However, there is a paucity of techniques for dual immunohistochemical (IHC) and mRNA in situ hybridization (ISH) detection. The recent development of improved non-radioactive detection systems and simplified ISH protocols has prompted us to develop a tyramide signal amplification method for sequential multi-label fluorescent ISH and IHC detection in either frozen or paraffin-embedded tissue sections. We used this method to examine the relationship between glial cell line-derived neurotrophic factor receptor α2 (GFRα2) mRNA expression and IHC localization of its co-receptor Ret in the trigeminal ganglion of postnatal Day 0 mice. We found that approximately 70% of Ret-immunoreactive neurons possessed GFRα2 mRNA and virtually all GFRα2-expressing neurons contained Ret-immunoreactive protein. Finally, we used paraformaldehyde-fixed, paraffin-embedded sections and a monoclonal antibody against neuron-specific nuclear antigen (NeuN) to demonstrate the neuronal specificity of GFRα2 mRNA expression in adult mouse brain. This multi-labeling technique should be applicable to a wide variety of tissues, antibodies, and probes, providing a relatively rapid and simple means to compare mRNA and protein localization.

Original languageEnglish
Pages (from-to)1369-1375
Number of pages7
JournalJournal of Histochemistry and Cytochemistry
Issue number10
StatePublished - 2000


  • Double labeling
  • Fluorescent in situ hybridization
  • Immunohistochemistry
  • Tyramide signal amplification


Dive into the research topics of 'Dual fluorescent in situ hybridization and immunohistochemical detection with tyramide signal amplication'. Together they form a unique fingerprint.

Cite this