Whole-Body Scanning PCR, a Tool for the Visualization of the In Vivo Biodistribution Pattern of Endogenous and Exogenous Oligonucleotides in Rodents

Methods Mol Biol. 2016:1372:99-111. doi: 10.1007/978-1-4939-3148-4_8.

Abstract

Characterizing the in vivo biodistribution pattern and relative expression levels of oligonucleotide-based molecules such as mRNA, miRNA, siRNA, and anti-miRNAs in animal models, could be a helpful first-step in the successful development of therapeutic oligonucleotides. Here we describe a simple procedure called "Whole-Body Scanning PCR" (WBS-PCR), which combines the power of PCR with that of imaging. WBS-PCR relies on 384 well-defined extractions across a mouse whole-body section followed by a single dilution step which renders the lysates compatible with various qPCR-based assays. The in vivo biodistribution maps are generated by deconvoluting the qPCR data and converting it into a TissueView compatible image file which can be overlaid with an image of the whole-body section used for extractions. WBS-PCR is a flexible platform that can be adapted to other detection systems and thereby further expand the use of this technology.

Keywords: Imaging; In vivo biodistribution; Mouse; QWBA; RT-qPCR; Rat; Therapeutic oligonucleotide s; miRNA; qPCR; siRNA.

MeSH terms

  • Animals
  • Cryoultramicrotomy / methods
  • Gene Expression*
  • Mice
  • MicroRNAs / genetics
  • Models, Animal
  • Molecular Imaging / methods*
  • Oligonucleotides / administration & dosage
  • Oligonucleotides / genetics*
  • Oligonucleotides / metabolism*
  • Polymerase Chain Reaction / methods*
  • RNA, Messenger / genetics
  • RNA, Ribosomal, 18S / genetics
  • RNA, Small Interfering / genetics
  • Rats
  • Real-Time Polymerase Chain Reaction / methods
  • Rodentia
  • Tissue Distribution
  • Whole Body Imaging / methods*

Substances

  • MicroRNAs
  • Oligonucleotides
  • RNA, Messenger
  • RNA, Ribosomal, 18S
  • RNA, Small Interfering