Fluorescence In Situ Hybridization (FISH)

Fluorescence in situ hybridization (FISH) is a molecular cytogenetic technique that uses fluorescently labelled DNA probes to bind specific chromosomal sequences, which are then visualised directly on chromosomes or in cell nuclei under a fluorescence microscope. By targeting defined loci rather than scanning the whole genome, FISH detects and localises specific deletions, duplications, rearrangements, and aneuploidies with high sensitivity, including in non-dividing (interphase) cells.

Definition

FISH is a technique in which a labelled nucleic acid probe is hybridized to complementary sequences fixed in place on a chromosome preparation or cell nucleus and detected by fluorescence, allowing specific genomic targets to be identified, counted, or localised.

Scope

This topic covers the principle of probe hybridization, the main probe types and their uses (locus-specific, centromeric, and whole-chromosome paint probes), and the application of FISH to metaphase and interphase cells. It is a methodological reference and does not provide clinical management guidance.

Core questions

• How does a labelled probe achieve specific, detectable binding to its chromosomal target?
• What distinguishes locus-specific, centromeric, and chromosome-paint probes?
• Why can FISH be applied to interphase as well as metaphase cells?
• What targeted abnormalities does FISH resolve that banding cannot?

Key concepts

• Nucleic acid hybridization
• Fluorescently labelled probe
• Locus-specific probe
• Centromeric (alpha-satellite) probe
• Whole-chromosome paint probe
• Interphase versus metaphase FISH
• Probe specificity and signal counting
• Microdeletion detection

Mechanisms

A DNA probe complementary to a target sequence is labelled with a fluorophore (directly or via a reporter molecule). The target chromosomal DNA on a slide and the probe are denatured to single strands and then allowed to anneal, so the probe hybridizes to its complementary sequence in place. After unbound probe is washed away, the bound signal is visualised by fluorescence microscopy. Different probe designs serve different purposes: locus-specific probes detect or confirm gains, losses, or rearrangements at a defined gene or region; centromeric probes count copies of a given chromosome (enumeration of aneuploidy); and whole-chromosome paint probes label an entire chromosome to characterise complex rearrangements. Because hybridization does not require dividing cells, FISH can be performed on interphase nuclei, extending analysis to tissues and samples in which metaphase preparations are difficult to obtain.

Clinical relevance

FISH is used to confirm or exclude specific microdeletion and microduplication conditions, to enumerate aneuploidies, and to detect defined rearrangements such as those characterising certain cancers. It complements karyotyping by adding targeted, high-sensitivity detection, including in interphase cells. This entry describes how FISH findings are generated; it is not a basis for individual diagnostic or treatment decisions.

Evidence & guidelines

FISH findings are described within the International System for Human Cytogenomic Nomenclature (ISCN), which includes notation for probe signals and hybridization results.

History

In situ hybridization was first developed with radioactive probes in the late 1960s. The shift to fluorescent detection, demonstrated quantitatively by Pinkel, Straume, and Gray in 1986, gave a faster, safer, and multicolour-capable method and launched molecular cytogenetics. Subsequent probe designs and multicolour approaches extended FISH from single-locus detection toward simultaneous analysis of many targets, bridging classical cytogenetics and molecular genetics.

Key figures

• Daniel Pinkel
• Joe W. Gray
• Michael Speicher
• Nigel Carter

Seminal works

• pinkel-1986
• speicher-carter-2005

Frequently asked questions

How is FISH different from karyotyping?

Karyotyping scans the whole genome at low resolution and reveals balanced rearrangements and ploidy, whereas FISH uses labelled probes to interrogate specific loci with high sensitivity, including in non-dividing interphase cells. They are complementary approaches.

Can FISH be performed without dividing cells?

Yes. Because hybridization does not depend on chromosome condensation, FISH can be applied to interphase nuclei, allowing analysis of samples from which metaphase chromosomes cannot easily be obtained.