Fluorescence in Situ Hybridization (FISH Test)

What is fluorescence in situ hybridization (FISH test)?

Fluorescence in situ hybridization (FISH) is a technique pathologists use in genetic testing to help diagnose diseases caused by chromosomal differences. Healthcare providers can also use FISH to detect gene changes that they can target to treat cancer. FISH uses proteins or chemical labels to “paint” parts of chromosomes or genes that pathologists look for to diagnose diseases. The labels look like colored lights under a microscope.

Your genes carry information on them in the form of DNA. DNA is like the instruction manual that tells your body how to work. Genes are organized on thread-like structures called chromosomes. If something is amiss in your instruction manual — like some words are deleted, repeated or moved around — it can give your cells the wrong instructions. Single genes or entire chromosomes of information can be missing (deletion), duplicated (amplification) or moved to a different place (translocation). These differences can cause changes in how your body works, or cause diseases like cancer.

What can FISH be used to test for?

Fluorescence in situ hybridization is most commonly used to test for:

• Large-scale gene mutations in cancer. Healthcare providers can use these gene mutations to diagnose or determine treatment.
• Chromosomal abnormalities before or after birth.
• Chromosomal abnormalities in an embryo during preimplantation genetic testing as a part of IVF (in vitro fertilization).

How does fluorescence in situ hybridization work?

If DNA is an instruction manual, a FISH test is a bit like highlighting important sections in special ink so a pathologist can find what they’re looking for. Scientists make this highlighter by marking strands of DNA with fluorescent labels. This is called the probe.

DNA comes in paired strands that match up with each other, like teeth on a zipper. Scientists separate those two strands in both their probe and in the sample they’re testing. They create the probe so it’s a match to one strand that has the DNA sequence — like a sentence on a page — that they’re looking for.

To perform the test, they mix their probes with the sample of DNA from body tissue or fluid (the target). If the probes find DNA with the sentence they’re looking for, they stick to it (hybridize). The labels highlight that section of DNA with bright colors. When a pathologist looks at it with a special microscope, they can see the labels lighting up, letting them know the information is there — and how much of it there is.

Gene changes detected by FISH

Pathologists can use FISH to test cells for gene mutations that are known to cause specific cancers or other conditions. It’s also sometimes used to confirm or clarify karyotyping findings. Specific changes that FISH can detect include:

• Amplifications. Amplification means that there are more copies of a gene than expected. Similar to finding extra chromosomes, scientists can use FISH to see if there are more copies of a specific gene than expected.
• Translocations/rearrangement. Translocation, or rearrangement, means that part of a gene or chromosome has moved to a different location. Scientists can use two different colored probes to find translocations. In a normal gene, the probes should appear next to each other. If they’re translocated, the colors will be separated.
• Deletions. Similar to translocations, scientists can use more than one probe to look for areas of DNA that are missing information. Some probes might stick to a section of the DNA in the sample and others don’t stick where they should. If the DNA isn’t labeled where it should be, some information is missing (deletion).

Providers can use FISH to detect gene changes that diagnose:

• Lymphoma.
• Acute myeloid leukemia, chronic myeloid leukemia and other forms of leukemia.
• Multiple myeloma.
• Bladder cancer.
• Glioma.
• Mesothelioma.
• HER2 amplification (most commonly found in breast cancer, but can also be found in stomach and lung cancer).
• MET amplifications (most commonly found in lung, stomach and esophageal cancer).
• MYCN amplification in neuroblastoma.
• ALK, RET and ROS1 rearrangement/fusion in non-small cell lung cancer. ALK, RET or ROS1 can rearrange and fuse with another gene, causing cancer.

How do I prepare for a FISH test?

Providers test a sample of body fluid or tissue for FISH analysis. Your preparation for the test depends on what type of sample your provider needs:

• For prenatal testing, your provider will perform an amniocentesis.
• For preimplantation testing (preimplantation genetic diagnosis/PGD or preimplantation genetic screening/PGS), your provider will test a few cells from embryos before implantation. This usually happens a few days after egg retrieval.
• If a provider thinks you or your child have a disease caused by a chromosomal abnormality or gene mutation, they can test for it with a blood test.
• To diagnose cancer or genetic mutations in tumor cells (molecular testing), you’ll need a tumor or bone marrow biopsy.
• A FISH urine (pee) test is sometimes used to diagnose or monitor bladder cancer.

Generally, the only one of these tests that might require preparation is a biopsy. Because many biopsies are done under anesthesia, you should follow your provider’s instructions on how to prepare.

How do pathologists perform fluorescence in situ hybridization?

For a FISH analysis, a pathologist or lab technician will:

1. Design the probe(s). This involves selecting sections of DNA that match the sequence they’re looking for. Then, they make small cuts in the DNA and add fluorescent labels.
2. Denature the probes and target. This means treating the probes and target so the two strands come apart.
3. Hybridization. When mixed with a sample of DNA from body tissue or fluid, probes will attach to sections of DNA that have the sequences they’re looking for.
4. Examine the results. They’ll use a fluorescent microscope to see what areas of the sample are highlighted with labels.

The pathologist will report the results to your provider.