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Related Concept Videos

FISH - Fluorescent In-situ Hybridization02:07

FISH - Fluorescent In-situ Hybridization

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,...
In-situ Hybridization02:31

In-situ Hybridization

In situ hybridization (ISH) is a technique used to detect and localize specific DNA or RNA molecules in cells, tissue, or tissue sections using a labeled probe. The technique was first used in 1969 for the investigation of nucleic acids. It is currently an essential tool in scientific research and clinical settings, especially for diagnostic purposes.
Types of probes and labels
A probe is a complementary strand of DNA or RNA that binds to corresponding nucleotide sequences in a cell. Many...

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Zebrafish Whole Mount High-Resolution Double Fluorescent In Situ Hybridization
12:31

Zebrafish Whole Mount High-Resolution Double Fluorescent In Situ Hybridization

Published on: March 25, 2009

Fluorescence in situ Hybridization (FISH).

Jane Bayani1, Jeremy A Squire

  • 1University of Toronto, Ontario, Canada.

Current Protocols in Cell Biology
|January 30, 2008
PubMed
Summary
This summary is machine-generated.

Fluorescence in situ Hybridization (FISH) is a molecular cytogenetic technique that visualizes specific DNA sequences. This method requires careful preparation of DNA probes and targets for accurate fluorescent microscopy interpretation.

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Area of Science:

  • Molecular Biology
  • Genetics
  • Cytogenetics

Background:

  • Fluorescence in situ Hybridization (FISH) is a powerful technique for visualizing specific DNA sequences within cells or tissues.
  • The method relies on the specific binding of a labeled DNA probe to a complementary target DNA sequence.

Purpose of the Study:

  • To outline the essential components and procedural steps involved in Fluorescence in situ Hybridization (FISH).
  • To highlight critical factors influencing the success and interpretation of FISH experiments.

Main Methods:

  • Preparation of DNA probes from various cloned sources (plasmids, cosmids, BACs) or whole-chromosome paints.
  • Labeling of DNA probes using direct or indirect methods with fluorochromes or haptens.
  • Preparation of target DNA from chromosome spreads or interphase nuclei, including those from paraffin-embedded tissues.
  • Denaturation of probe and target DNA, followed by hybridization, post-hybridization washes, and antibody incubations.
  • Visualization of results using fluorescent microscopy.

Main Results:

  • Successful FISH experiments depend on high-quality starting materials, efficient hybridization, and optimized stringency of washes and antibody detection.
  • Various labeling strategies (nick translation, PCR labeling) and probe sources can be employed based on experimental needs.
  • FISH can be applied to different target DNA preparations, including metaphase chromosomes, interphase nuclei, and tissue sections.

Conclusions:

  • FISH is a versatile technique with critical steps requiring meticulous execution for reliable results.
  • The quality of reagents and adherence to optimized protocols are paramount for accurate DNA sequence visualization and interpretation in FISH assays.