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Immunohistochemistry (IHC)

This page was reviewed under our medical and editorial policy by

Bradford Tan, MD, Chair, CTCA Department of Pathology and Laboratory Medicine.

This page was reviewed on September 25, 2022.

Immunohistochemistry (IHC) is a diagnostic technique conducted in a laboratory. It involves performing special tests on your biopsy tissue sample to help diagnose disease more precisely. IHC helps your care team distinguish between different types and subtypes of a disease by identifying specific molecules within cells, known as markers.

IHC is particularly useful in diagnosing certain types of cancer, because your care team often needs a wealth of information about the disease to help determine treatment, such as:

  • Where the cancer started
  • The type of cell it started in
  • Whether it’s likely to grow slowly or quickly

IHC may help provide these answers.

Before performing an IHC test, your care team must collect a tissue sample. The procedure to remove an abnormal or cancerous specimen is called a biopsy. Methods range from needle biopsies, which involve inserting a needle into the tumor, to excisional biopsies, which remove the entire tumor. The method used depends on individual circumstances, including the type of cancer and its location.

All biopsy types involve removing bodily tissue and taking it to a laboratory, where a pathologist who specializes in diagnosing diseases will examine the sample under a microscope. To determine whether or not the specimen is cancerous, the pathologist usually only needs to examine the cells and perform standard tests called stains, which use colored substances (dyes) to highlight cancer cells.

It can be difficult to differentiate cancer cells based only on their appearance and routine tests—and when these methods aren’t enough to make a diagnosis, your care team may turn to IHC.

IHC reveals more than a standard biopsy test. It enables pathologists to determine the exact type and subtype of cancer by looking for unique markers within cancer cells. Markers are identified using antibodies, a type of protein that locates and binds with antigens (which are the markers found on cells). Antigens are the lock, and antibodies are the key—unique antibodies fit perfectly within their respective antigens.

More than 400 unique antibodies can be used in IHC tests. When an antibody recognizes and binds to corresponding antigen in the sample, a colored dye is activated, indicating to the pathologist that the sample contains a particular type of cell that may help determine the exact type of cancer.

Why IHC is done

There are many different circumstances in which IHC tests can be helpful, but these tests aren’t often part of the standard diagnostic process for most cancer types. For the most part, IHC tests are only ordered after a pathologist has reviewed the results of more routine tests.

Pathologists can diagnose most cancers just by looking at a biopsy sample under a microscope and using stains. However, if the analysis is more complex and requires checking for more specific characteristics to make a diagnosis, the pathologist can turn to IHC tests.

Your doctor and a pathologist can recommend whether or not IHC is necessary on a case-by-case basis.

However, IHC tests are routinely used to help diagnose most types of breast cancer, including:

In patients with breast cancer, IHC is used to test for:

Hormone receptor status: IHC tests can detect the presence or absence of hormone receptors on breast cancer cells. This knowledge informs how the cancer may be treated, as breast cancers that carry these receptors can be treated with hormone therapy drugs.

HER2 status: IHC tests can check for HER2 receptors to determine whether breast cancer is HER2-positive or HER2-negative. If these receptors are found, the cancer is HER2-positive, which indicates that it’s likely to be fast-growing and can be treated by targeted therapy drugs that block the effects of the HER2 proteins responsible for fueling the cancer’s growth.

Another routine use of IHC testing is for Lynch syndrome, an inherited condition that can cause cancer or increase one’s risk of developing cancer. Lynch syndrome is most often linked to colorectal cancer, but it can also raise the risk of cancer in the uterus, stomach, liver, kidney and brain. Doctors may recommend using IHC to check for markers of Lynch syndrome in women who develop endometrial cancer, or anyone diagnosed with colorectal cancer before age 70.

For other cancer types, IHC testing may be warranted under certain circumstances. These include:

Gastrointestinal cancer: Some gastric cancers are linked to autoimmune diseases or bacterial strains, such as H. pylori. IHC tests can help inform treatment by determining whether these or other factors may be responsible for causing the cancer. IHC tests can also help differentiate between types of gastrointestinal cancer.

Prostate cancer: Some prostate cancers are difficult to diagnose using standard tests. Numerous antibodies can be used in IHC tests to learn more about the cancer, although performing these tests isn’t always necessary. IHC tests are more likely to be used if standard tests reveal mixed findings.

Lung cancer: After other tests confirm a lung cancer diagnosis, IHC tests may be necessary to define the type of lung cancer. For example, a pathologist will often need to use IHC to check for specific antigens to establish a case of non-small cell lung cancer.

Lymphoma: It can be difficult for a pathologist to recognize whether swollen lymph nodes are caused by an infection or cancer such as lymphoma. With IHC tests, the pathologist can test the white blood cells causing the swelling to see whether they carry cancer-indicating antigens. IHC tests may also help distinguish between different lymphoma types.

Other cancers that may sometimes need IHC testing include those of the female reproductive system, bladder and kidney. However, many other circumstances warrant using the IHC method. The decision to use IHC tests is based on careful consideration of individual factors.

What to expect and how to prepare

This type of testing is an additional step performed on your biopsy sample, so you don’t need to prepare specifically for IHC. You’ll get ready for your biopsy, however.

Biopsy types vary significantly, ranging from a quick procedure performed in your doctor’s office to an invasive surgery that may come with risks. Your care team will provide instructions to help you prepare and let you know what to expect. For example, if you need general anesthesia, you may be told not to eat or drink for a certain period of time leading up to the biopsy and may need someone to drive you home afterward.

Before or after a biopsy, your care team can inform you as to whether IHC testing is necessary or important to help them form a more precise diagnosis and treatment plan. If you’re not sure, ask your doctor which lab tests were ordered. You may also want to ask whether your insurance covers these tests. Insurance may cover IHC testing for certain reasons but not for others.

How IHC is performed

After a biopsy procedure, the tissue is taken to a laboratory for a pathologist to examine the sample with and without a microscope. The biopsy specimen can be sliced into thin pieces for microscopic examination. The slices are typically also dipped in colorful stains (dyes) to help the pathologist observe the cells and differentiate between cell types.

For an IHC test, the pathologist starts by considering the type of cancer suggested by previous tests and then selects an antibody (or several) that might help answer the remaining questions.

For example, if the sample is known to contain breast cancer cells but the pathologist needs to find out if the cells are hormone receptor-positive, he or she will use an estrogen or progesterone antibody that is known to bind with these hormone receptors. Along with the antibody, the pathologist may add chemicals or dyes to the solution. The dyes change the color of cells that contain hormone receptors, making them easily identifiable. The pathologist will then dip a slice of the sample into the antibody solution and place it under the microscope to examine the findings.

Understanding the results

Once the analysis is complete, a pathology report will be prepared to summarize the diagnosis and other relevant information. It may take about two to 10 days for results to be finalized and presented to you after a biopsy procedure. The diagnostic process can be longer or shorter, depending on the number of tests performed and their complexity. IHC tests typically only take one day longer than routine tests. More complicated analyses, such as flow cytometry, tend to delay the process further.

Once you receive your pathology report, look it over and take note of any questions or concerns you may have. Your doctor may go through the report in detail with you.

Benefits and risks of IHC

The benefit of IHC testing is that, when necessary, it allows your care team to gain a more precise understanding of the cancer and how best to treat it. However, these tests aren’t 100 percent accurate. Misdiagnosis of any sort is harmful, with false-positives resulting in unnecessary treatment and trauma, and false-negatives leading to potentially detrimental treatment delays.

The rate of false-positives and false-negatives vary significantly with an IHC. Numerous factors may influence the accuracy of these tests, including:

  • Technology, equipment and materials used
  • Storage and handling of the sample
  • Errors made throughout the process

IHC test kits are regulated by the U.S. Food and Drug Administration. All tests must present data on their accuracy in repeated attempts and against other methods. Pathologists are also encouraged to regularly validate their own IHC systems to ensure that they are reporting accurate results.

While a cancer misdiagnosis can occur, systems in place are designed to help avoid this, such as requiring a second pathologist to review findings (in most accredited labs) and having a team of doctors and pathologists work together to interpret the results.

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