Chromogen detection of microRNA in frozen clinical tissue samples using LNA™ probe technology

Methods Mol Biol. 2014:1211:77-84. doi: 10.1007/978-1-4939-1459-3_7.

Abstract

Specific chromogen- and fluorescence-based detection of microRNA by in situ hybridization (ISH) in formalin-fixed and paraffin-embedded (FFPE) tissue sections has been facilitated by locked nucleic acid (LNA)-based probe technology and can be performed within a single working day. In the current method paper, we present a similar simple 1-day ISH method developed for cryostat sections obtained from clinical cryo-embedded tissue samples. The presented chromogen-based ISH method does not involve proteolytic pretreatment, which is mandatory for FFPE sections, but still retains a sensitivity level similar to that obtained in FFPE sections. The LNA-based ISH method is not only applicable in situations where only access to cryo-embedded material is possible, but it also has a potential use if combining microRNA ISH with immunohistochemistry in double fluorescence staining with antibodies not being compatible with proteolytic predigestion.

MeSH terms

  • Cryopreservation
  • Fluorescent Dyes / analysis
  • Formaldehyde / chemistry
  • Humans
  • Immunohistochemistry
  • In Situ Hybridization / economics
  • In Situ Hybridization / methods*
  • MicroRNAs / analysis*
  • MicroRNAs / genetics
  • Oligonucleotides / analysis*
  • Oligonucleotides / genetics
  • Paraffin Embedding / economics
  • Paraffin Embedding / methods*
  • Tissue Fixation / economics
  • Tissue Fixation / methods*

Substances

  • Fluorescent Dyes
  • MicroRNAs
  • Oligonucleotides
  • locked nucleic acid
  • Formaldehyde