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FISH - Fluorescent In-situ Hybridization

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Fluorescence in situ hybridization, or FISH, was developed in the early 1980s and has quickly become one of the most widely used techniques in cytogenetics. Labeled probes are used to bind complementary DNA or RNA sequences on a chromosome or in a region within a cell. Earlier, the probes could only be obtained by cloning or reverse transcription of a DNA template. Currently, the probe oligonucleotides can be synthesized synthetically. Additionally, with the advancement of optical techniques,...

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Updated: Jun 13, 2025

Robust 3D DNA FISH Using Directly Labeled Probes

Robust 3D DNA FISH Using Directly Labeled Probes

Published on: August 15, 2013

Image Analysis Protocol for DNA/RNA/Immunofluorescence (IF)-seqFISH Data.

Hiroaki Ohishi¹, Hiroshi Ochiai²

¹Division of Gene Expression Dynamics, Medical Institute of Bioregulation, Kyushu University, Fukuoka, Japan.

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

|September 16, 2024

Summary

This study details imaging-based spatial multi-omics, focusing on sequential fluorescence in situ hybridization (seqFISH). It provides a practical image analysis protocol for observing gene transcription and protein localization within cells and tissues.

Keywords:

Higher-order genomic structure Imaging Transcription seqFISH

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Last Updated: Jun 13, 2025

Robust 3D DNA FISH Using Directly Labeled Probes

Robust 3D DNA FISH Using Directly Labeled Probes

Published on: August 15, 2013

Single Cell Analysis Of Transcriptionally Active Alleles By Single Molecule FISH

Single Cell Analysis Of Transcriptionally Active Alleles By Single Molecule FISH

Published on: September 20, 2020

Workflow for High-content, Individual Cell Quantification of Fluorescent Markers from Universal Microscope Data, Supported by Open Source Software

Workflow for High-content, Individual Cell Quantification of Fluorescent Markers from Universal Microscope Data, Supported by Open Source Software

Published on: December 16, 2014

Area of Science:

Molecular Biology
Genomics
Cell Biology

Background:

Imaging-based spatial multi-omics enable analysis of genomic structures, gene transcription, and protein localization at the single-allele level.
These technologies offer detailed observations of biological phenomena, including intracellular transcription machinery.

Purpose of the Study:

To detail the principles of imaging-based spatial multi-omics technologies.
To provide a comprehensive protocol for sequential fluorescence in situ hybridization (seqFISH) image analysis.
To enable practical application through accessible code.

Main Methods:

Focus on DNA/RNA/immunofluorescence (IF) sequential fluorescence in situ hybridization (seqFISH).
Detailed step-by-step protocol for image analysis: preprocessing, spot detection, and data visualization.
Provision of complete Jupyter Notebook codes for practical application.

Main Results:

Demonstration of principles for spatial multi-omics analysis.
A functional protocol for seqFISH image analysis is presented.
Open-access code facilitates reproducible research.

Conclusions:

Imaging-based spatial multi-omics, particularly seqFISH, are powerful tools for biological research.
The provided protocol and code enhance the accessibility and application of these advanced techniques.
Facilitates detailed observation of molecular events within cellular and tissue contexts.