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Rapid Cold Fixation of Mouse Colon (Col'RFix) Enables High-Resolution Mass Spectrometry Imaging.

Sabina H Skov^1,2, Henrik M Jensen², Ole N Jensen¹

¹Department of Biochemistry and Molecular Biology, University of Southern Denmark, DK-5230 Odense M, Denmark.

Analytical Chemistry

|April 13, 2026

Summary

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A new rapid fixation method, Col

Area of Science:

Mass spectrometry imaging
Tissue preparation
Molecular imaging

Background:

Cellular-scale MALDI MSI demands tissue prep preserving morphology and molecular integrity.
Mouse colon tissue is challenging due to fragility and fixation artifacts.

Purpose of the Study:

Introduce Col'RFix, a novel rapid fixation workflow for mouse colon.
Optimize tissue preparation for high-resolution MALDI MSI of the colon.

Main Methods:

Col'RFix uses cold PFA fixation and agarose embedding for mechanical support.
Protocol validated for morphology, analyte diffusion, molecular coverage, and spatial localization.
Evaluated across different pixel sizes (5-20 μm), MSI modalities, and ion modes.

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Related Experiment Videos

Last Updated: Apr 14, 2026

Performing Colonoscopic-Guided Pinch Biopsies in Mice and Evaluating Subsequent Tissue Changes

Performing Colonoscopic-Guided Pinch Biopsies in Mice and Evaluating Subsequent Tissue Changes

Published on: February 5, 2021

Cryosectioning Method for Microdissection of Murine Colonic Mucosa

Cryosectioning Method for Microdissection of Murine Colonic Mucosa

Published on: July 12, 2015

Three-dimensional Quantification of Intestinal Mucus Using Whole-mount Tissue Imaging

Three-dimensional Quantification of Intestinal Mucus Using Whole-mount Tissue Imaging

Published on: September 12, 2025

Main Results:

Col'RFix enables reproducible imaging at 5 μm pixel size with minimal delocalization.
Preserves fine colon morphology, allowing correlation with histological details (H&E staining).
Maintains largely conserved molecular profiles compared to unfixed tissue, with some signal attenuation.

Conclusions:

Col'RFix is a reproducible workflow for compartment-resolved molecular mapping of mouse colon.
Facilitates high-resolution spatial lipidomics of colon tissue.
Enables precise correlation of molecular features with histological structures.