Transglutaminase-mediated in situ hybridization (transish) for mRNA detection in mammalian tissues

Katsuyuki Miyawaki, Sumihare Noji, Noriho Kamiya

Research output: Chapter in Book/Report/Conference proceedingChapter


In the most commonly used in situ hybridization (ISH) procedure, a hapten-labeled antisense nucleic acid (e.g., RNA) probe is employed to hybridize a target mRNA in tissue sections. The hapten-labeled RNA is then detected by a highly specific hapten-antibody interaction; however, it requires laborious immunostaining steps for spatial coloring on tissue sections. To simplify ISH-based mRNA detection systems, we created a new RNA-(enzyme) n conjugate for sensitive detection of mRNA in tissue sections. In the present simple, antibody-free ISH protocol, an antisense RNA probe of interest is first modified with a specific dipeptide substrate of microbial transglutaminase (MTG) by in vitro transcription. Alkaline phosphatase from a hyperthermophile is then covalently linked to the substrate-labeled RNA by MTG-catalyzed sitespecific conjugation. A robust, multi-enzyme-labeled RNA probe enables the direct labeling of a target mRNA in tissue sections with signaling enzymes under harsh hybridization conditions, leading to one-step signal amplification after hybridization. The application of the new trans glutaminase-mediated ISH (TransISH) strategy to mRNA detection in mammalian tissues was demonstrated.

Original languageEnglish
Title of host publicationIn Situ Hybridization Methods
PublisherSpringer New York
Number of pages10
ISBN (Electronic)9781493923038
ISBN (Print)9781493923021
Publication statusPublished - Feb 18 2015

All Science Journal Classification (ASJC) codes

  • General Medicine
  • General Neuroscience
  • General Agricultural and Biological Sciences


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