Fluorescence In Situ Hybridization (FISH) in Cytology

Fluorescence in situ hybridization (FISH) uses fluorescently labelled DNA probes that bind to complementary sequences in cell nuclei, making specific chromosomal regions or genes visible under a fluorescence microscope. Applied to cytologic specimens, it can detect aneuploidy, gene amplifications, deletions, and rearrangements directly in intact cells, adding a cytogenetic dimension to morphologic assessment.

Definition

FISH is an ancillary cytogenetic technique in which fluorescently labelled nucleic-acid probes hybridize to specific DNA targets within cells on a cytologic preparation, allowing visualization of chromosomal copy-number changes and structural rearrangements.

Scope

The entry covers the principle of probe hybridization on cytologic preparations, the kinds of genomic abnormalities FISH can resolve, and its established cytologic applications such as multitarget assays in urinary and bronchial specimens. It is a methodological reference and does not provide testing or interpretation protocols.

Core questions

• Which chromosomal or gene-level abnormalities can FISH resolve in a given cytologic specimen?
• How do probe design and signal-enumeration criteria determine a positive result?
• How does FISH complement cytologic morphology in equivocal or atypical samples?

Key concepts

• Sequence-specific probe hybridization
• Centromeric, locus-specific, and break-apart probes
• Aneuploidy and copy-number detection
• Multitarget probe panels (e.g., UroVysion)
• Signal enumeration and scoring thresholds
• Application to intact cells on slides

Mechanisms

Denatured single-stranded probe DNA, carried by a fluorophore, anneals to its complementary target sequence within the denatured nuclear DNA of cells fixed on a slide; after washing away unbound probe, the bound signals are counted under fluorescence microscopy. Centromeric probes report chromosome copy number, locus-specific probes detect amplification or deletion of a region, and dual-colour break-apart probes reveal rearrangements by the separation of paired signals. Multitarget panels combine several probes so that aneuploidy patterns characteristic of malignancy can be recognized in cells that look ambiguous morphologically.

Clinical relevance

FISH adds objective cytogenetic information to cytology, for example helping to evaluate atypical urothelial samples or to assess pulmonary cytology, and it is used as an adjunct rather than a replacement for morphologic review. This entry describes how the method generates information; specific assay selection and interpretation are laboratory and clinical decisions and are not individualized advice.

Evidence & guidelines

Paired-sample studies of the multitarget UroVysion assay in voided urine have compared its performance with conventional cytology for detecting urothelial carcinoma, reporting complementary strengths and limitations (Lavery et al., 2017; Dimashkieh et al., 2013). Multitarget FISH has likewise been evaluated on bronchial cytology specimens for the detection of lung cancer (Zhai et al., 2015).

History

In situ hybridization was first developed for tissue and chromosome preparations; the substitution of fluorescent labels for radioactive ones made multicolour, multi-probe assays practical and allowed the technique to be applied directly to cytologic slides. Multitarget urinary panels became one of the most widely studied cytologic applications.

Debates

How should FISH and conventional cytology be combined in atypical urinary samples?

Studies show FISH and cytology have complementary sensitivity and specificity, raising the question of whether FISH is best used as a reflex test, an adjunct to morphology, or a triage tool, with no single approach universally adopted.

Seminal works

• dimashkieh-2013
• lavery-2017

Frequently asked questions

What kinds of abnormalities can FISH detect in cytologic samples?

FISH can reveal chromosomal aneuploidy, gene amplifications and deletions, and structural rearrangements by counting fluorescent probe signals in individual cell nuclei on the slide.

Does FISH replace conventional cytology?

No; it is used as an adjunct. FISH and morphologic cytology have complementary strengths, and FISH adds cytogenetic information rather than substituting for visual assessment of the cells.