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Protocol to characterize mouse intestinal epithelial cell lineage using Opal multiplex immunofluorescence.

Anxo Martinez-Ordoñez1, Hiroto Kinoshita1, Tania Cid-Diaz1

  • 1Department of Pathology and Laboratory Medicine, Weill Cornell Medicine, New York, NY 10065, USA.

STAR Protocols
|September 12, 2024
PubMed
Summary

This study presents a protocol for using Opal multiplex immunofluorescence (OMI) to characterize mouse intestinal epithelial cell lineage. The method optimizes signal detection for multiple targets in genetically engineered mouse models.

Keywords:
CancerIn Situ HybridizationMicroscopyModel OrganismsMolecular BiologyMolecular/Chemical ProbesStem Cells

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Area of Science:

  • Immunohistochemistry and cell biology
  • Gastrointestinal research
  • Mouse model studies

Background:

  • Opal multiplex immunofluorescence (OMI) is a powerful tool for analyzing complex biological processes in tissues.
  • Characterizing intestinal epithelial cell lineage in mouse models is crucial for understanding gastrointestinal diseases.
  • Detecting multiple target molecules simultaneously in intestinal tissues using OMI presents technical challenges.

Purpose of the Study:

  • To present a detailed protocol for characterizing mouse intestinal epithelial cell lineage using OMI.
  • To optimize OMI panel design and signal detection for mouse intestinal tissues.
  • To provide a method for quantitative evaluation of OMI images in this context.

Main Methods:

  • Protocol development for processing mouse small intestine and colonic tissues.
  • Design and optimization of OMI panels for multi-target detection in mouse intestines.
  • Quantitative image analysis procedures for OMI data.

Main Results:

  • A standardized protocol for OMI application in mouse intestinal tissues was established.
  • Optimized OMI panels enable robust detection of multiple intestinal epithelial cell markers.
  • Quantitative evaluation methods allow for reliable assessment of cell lineage markers.

Conclusions:

  • The developed OMI protocol is effective for characterizing mouse intestinal epithelial cell lineage.
  • This method enhances the study of complex cellular processes in genetically engineered mouse models.
  • The protocol facilitates detailed spatial and quantitative analysis of intestinal tissue architecture.