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Updated: May 2, 2026

High-plex Imaging using Spectral Confocal Microscopy to Minimize Non-specific Tissue Fluorescence

High-plex Imaging using Spectral Confocal Microscopy to Minimize Non-specific Tissue Fluorescence

Published on: October 28, 2025

Current Methods in CODEX High-Multiplex Microscopy to Describe Spatial Relationships in the Tissue Microenvironment.

Joseph Nichols¹, Amina Nurmammadova^1,2, Yannick Palmowski¹

¹Department of Pathology and Neuropathology, University Hospital and Comprehensive Cancer Center Tübingen, Tübingen, Germany.

Methods in Molecular Biology (Clifton, N.J.)

|April 30, 2026

Summary

This summary is machine-generated.

High-multiplex microscopy using CO-Detection by indEXing (CODEX) with oligonucleotide-conjugated antibodies enhances tissue and cellular level resolution for studying disease mechanisms. This method maps cell types, neighborhoods, and functional states for improved diagnostics and treatments, especially in cancer.

Keywords:

CODEX Cancer immunology High-multiplex microscopy Personalized therapy Tissue microenvironment

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Visualization, Quantification, and Mapping of Immune Cell Populations in the Tumor Microenvironment

Visualization, Quantification, and Mapping of Immune Cell Populations in the Tumor Microenvironment

Published on: March 25, 2020

Area of Science:

Biomedical Engineering
Molecular Biology
Pathology

Background:

Studying disease mechanisms at the cellular and tissue levels requires high-resolution imaging techniques.
Existing methods may lack the multiplexing capability to analyze complex cellular interactions within tissues.

Purpose of the Study:

To introduce and detail the CO-Detection by indEXing (CODEX) method for high-multiplex tissue imaging.
To enable detailed spatial analysis of cell types, neighborhoods, and functional states within tissues.

Main Methods:

Antibodies are conjugated to unique oligonucleotides.
Conjugates are optimized for formalin-fixed paraffin-embedded (FFPE) or fresh-frozen (FF) tissues.
CODEX involves a multicycle reaction with fluorescently labeled oligonucleotides for image capture, followed by image processing for spatial analysis.

Main Results:

CODEX allows for high-multiplex imaging, analyzing up to 100 unique targets simultaneously.
The method facilitates the identification of cell types, cellular neighborhoods, and their spatial relationships.
Image processing enables cell type and neighborhood mapping for spatial analysis.

Conclusions:

CODEX significantly advances basic antibody research and translational pathology.
This high-resolution spatial analysis method refines clinical treatments by providing bioinformatic support for disease diagnosis, particularly in cancer.
The technique offers a powerful tool for understanding complex biological systems at the microscopic level.