Fluorescence in situ hybridisation (FISH)

What it is

Fluorescence in situ hybridisation (FISH) is a lab test that uses fluorescent DNA probes to detect specific chromosome or gene changes in cells, and one key practical fact is that it can often be performed on formalin-fixed paraffin-embedded tissue as well as blood, bone marrow, or fresh samples. In plain language, FISH looks for whether a chosen DNA sequence is present, missing, duplicated, amplified, or rearranged, and shows the result as coloured signals under a fluorescence microscope. It is widely used in cancer diagnostics, especially when doctors need to confirm a gene fusion, gene amplification, deletion, or chromosome copy-number change.

FISH is a targeted test. That means it does not scan the whole genome in the way some newer sequencing or microarray methods can. Instead, it answers a focused question such as whether HER2 is amplified in breast cancer, whether BCR::ABL1 is present in chronic myeloid leukaemia, or whether a lymphoma has a specific rearrangement. Because of this, FISH is often used alongside histopathology, immunohistochemistry, karyotyping, PCR, or next-generation sequencing rather than replacing them.

Common sample types include:

Sample type	Common use
Tumour tissue on slides	Solid tumours such as breast, lung, sarcoma
Blood or bone marrow	Leukaemia and lymphoma
Cell preparations/cultures	Cytogenetic work-up
FFPE tissue blocks	Retrospective or routine pathology testing

How it works

FISH uses short pieces of labelled DNA called probes. Each probe is designed to bind, or hybridise, to a specific DNA sequence in the cell nucleus or on a chromosome. After the sample is prepared on a slide, the DNA is denatured so the strands separate. The fluorescent probe is then added and allowed to bind to its matching target. Under a fluorescence microscope, the bound probes appear as coloured dots or patterns.

The signal pattern tells the laboratory what kind of abnormality may be present. For example:

1. Fusion probes can show that two genes have joined together because of a translocation.
2. Break-apart probes can show that a gene region has split, suggesting a rearrangement.
3. Enumeration probes can count chromosome copies, such as extra copies of chromosome 8.
4. Amplification probes can show many extra copies of a gene, such as HER2.
5. Deletion probes can show loss of a specific region.

FISH can be done on dividing cells or on interphase nuclei, which is one reason it is faster and more practical than conventional karyotyping in many settings. It also preserves spatial information, meaning the lab can see the signal inside individual cells rather than only reading extracted DNA.

Evidence and uses

FISH is an established clinical test in genetics and oncology. In cancer care, it helps with diagnosis, prognosis, and treatment selection. In haematologic cancers, FISH is commonly used to detect recurrent chromosome abnormalities and fusion genes. In solid tumours, it is often used to confirm gene amplification or rearrangement when that result affects therapy.

Examples in cancer include:

Cancer setting	Example target	Why it matters
Breast cancer	HER2 amplification	Helps decide anti-HER2 treatment
Chronic myeloid leukaemia	BCR::ABL1 fusion	Confirms a defining genetic abnormality
Acute promyelocytic leukaemia	PML::RARA	Urgent diagnostic and treatment relevance
Lung cancer	ALK, ROS1, RET rearrangements	May guide targeted therapy
Lymphoma	MYC, BCL2, BCL6 rearrangements	Helps classify disease and estimate risk
Plasma cell disorders	Common cytogenetic abnormalities	Supports risk stratification

FISH is also used outside cancer, including some constitutional genetic disorders and prenatal or postnatal chromosome studies. However, in several non-cancer settings it has been partly replaced by chromosomal microarray, QF-PCR, or sequencing-based methods because those tests can survey more of the genome at once.

In India, FISH is available in many tertiary cancer centres and reference laboratories, especially for breast cancer, lung cancer, leukaemia, lymphoma, and sarcoma work-ups. The exact panel used depends on the tumour type, pathology findings, and whether the result will change management. Because FISH is a targeted and relatively labour-intensive test, clinicians usually order it only when there is a clear clinical question.

Diagnosis / how it's measured

A FISH result is not simply "positive" or "negative" in the everyday sense. The laboratory counts fluorescent signals in a defined number of cells and interprets the pattern using validated cut-offs for that assay. The report usually includes the probe used, specimen type, number of cells scored, signal pattern, and interpretation.

Important points about measurement and reporting:

• Specimen quality matters. Poor fixation, low tumour content, or damaged nuclei can reduce accuracy.
• Cut-offs are assay-specific. A positive threshold for one probe set is not automatically the same for another.
• Tumour heterogeneity can affect results. Some parts of a tumour may carry the abnormality while others do not.
• Context matters. FISH findings are interpreted with pathology, imaging, blood counts, morphology, and other molecular tests.

FISH has strengths and limits compared with other methods:

Test	Main strength	Main limitation
FISH	Targeted, visual, works on many specimen types	Only detects what the probe is designed to find
Karyotype	Genome-wide view of large chromosome changes	Needs dividing cells, lower resolution
PCR/RT-PCR	Very sensitive for known targets	Usually narrower and less visual
NGS	Broad molecular profiling	More complex, costlier, may take longer
Microarray	Genome-wide copy-number analysis	Does not detect balanced rearrangements well

When to see a clinician

Patients do not usually decide on FISH testing on their own. A clinician may suggest it after a biopsy, bone marrow test, or pathology review if the result could confirm a diagnosis or guide treatment. Ask your oncologist or pathologist why the test is being ordered, what abnormality is being looked for, and whether the result could change treatment choices.

You should also ask for help interpreting the report if you see terms such as amplification, rearrangement, translocation, deletion, monosomy, polysomy, or break-apart signal. These terms are technical, and their meaning depends on the cancer type and the exact probe used.

Limitations and open questions

FISH is highly useful, but it is not a complete genetic test. It can miss abnormalities outside the targeted region, unexpected variants, or very complex genomic changes. A normal FISH result does not always rule out cancer or rule out a molecular abnormality detectable by another method.

Another limitation is that different laboratories may use different probe sets, scoring methods, and reporting formats, although accredited labs follow validation and quality-control standards. Borderline or equivocal results can occur, especially when tissue quality is poor or the signal pattern is difficult to interpret.

As cancer testing evolves, FISH remains important but is increasingly used as part of a combined diagnostic strategy. In some cancers, next-generation sequencing or broader genomic panels may provide more information. Even so, FISH still has a strong role when a fast, targeted answer is needed, when tissue is limited, or when a specific rearrangement or amplification is best seen directly in cells.