Crohn's disease: Failure of a proprietary fluorescent in situ hybridization assay to detect M. avium subspecies paratuberculosis in archived frozen intestine from patients with Crohn's disease.

Robert J. Greenstein, Liya Su, Peter S. Fam, Brooke Gurland, Paul Endres, Sheldon T. Brown

Research output: Contribution to journalArticlepeer-review

Abstract

Objectives: Although controversial, there is increasing concern that Crohn's disease may be a zoonotic infectious disease consequent to a mycobacterial infection. The most plausible candidate is M. avium subspecies paratuberculosis (MAP) that is unequivocally responsible for Johne's disease in ruminants. The purpose of this study was to evaluate a proprietary (Affymetrix™ RNA view®) fluorescent in situ hybridization (FISH) assay for MAP RNA. Non-identifiable intestine from patients with documented Crohn's disease was assayed according to the manufacturer's instructions and with suggested modifications. Probes were custom designed for MAP and human β-actin (as the eukaryotic housekeeping gene) from published genomes. Results: Repetitively, false positive signal was observed in our "No-Probe" negative control. Attempts were made to correct this according to the manufacturer's suggestions (by modifying wash solutions, using recommended hydrochloric acid titration and different fluorescent filters). None prevented false positive signal in the "No-Probe" control. It is concluded that when performed according to manufactures instruction and with multiple variations on the manufactures recommended suggestions to correct for false positive signal, that the Affymetrix™ RNA view® cannot be used to detect MAP in pre-frozen resected intestine of humans with Crohn's disease.

Original languageEnglish
Article number96
JournalBMC Research Notes
Volume13
Issue number1
DOIs
StatePublished - 24 Feb 2020

Keywords

  • Crohn disease
  • In situ hybridization
  • Johne disease
  • Mycobacteria
  • Mycobacterium avium subspecies paratuberculosis

Fingerprint

Dive into the research topics of 'Crohn's disease: Failure of a proprietary fluorescent in situ hybridization assay to detect M. avium subspecies paratuberculosis in archived frozen intestine from patients with Crohn's disease.'. Together they form a unique fingerprint.

Cite this