Immunohistochemistry is a technique used to determine the presence and level of specific cellular proteins. IHC measures proteinOne of the four basic types of biomolecule. Proteins are polymers made up of strings of amino acids. Proteins serve many functions in organisms including transport of molecules, structure, cell adhesion and as signaling molecules such as hormones. Many transcription factors, including p53 and Rb are proteins. expression using specially labeled antibodiesAntibodies are proteins produced by a type of white blood cell (B cells or B lymphocytes). Antibodies are able to stick very tightly to specific targets. Antibodies are currently being used as anti-cancer drugs (i.e. Herceptin). that can bind to the proteins of interest. The antibodyAntibody molecules tightly bind to their targets (usually proteins) with great specificity. Antibody binding alerts the immune system to target the bound molecule for destruction. Specific antibodies can be made in the laboratory. is mixed with the cellular components of the tumor. After a set amount of time, the mixture is rinsed and only those antibodies attached to their protein targets will remain. The presence of the antibodies can be detected by viewing the sample under a microscope because areas containing bound antibodies will appear a different color than areas lacking antibodies. Samples with more protein will bind more antibody and therefore appear darker. This allows the test to reveal not only whether a protein is present but also the relative amount of the protein. Test results are based on the strength of the staining and the percent of cells stained.1, 2, 3

IHC is a commonly used technique because it is inexpensive to perform, doesn't require special equipment, and generally quite accurate.2, 1, 3

As an example, three proteins of particular interest for breast cancer are HER2, the estrogen receptorA protein located within cells that binds estrogen. The protein:estrogen complex then binds to DNA to increase the transcription of certain genes, and therefore is a type of transcription factor. The estrogen receptor is the target of the anticancer drug tamoxifen. By binding to the estrogen receptor tamoxifen blocks the growth promoting effects of the hormone. May be abbreviated as ER. (ERThe endoplasmic reticulum. A large subcellular network of membranes within the cell. The ER membrane, an extension of the outer nuclear membrane, has two structurally and functionally distinct regions: Rough ER (RER) and Smooth ER (SER). The RER ( the portion closer to the nucleus) has a 'rough' appearance due to ribosomes attached to the surface of the membrane. Ribosomes attached to the ER are producing proteins that are to be released from the cell. Smooth ER (SER) appears smooth and is devoid of ribosomes. SER functions include the detoxification of drugs, the production of steroid hormones and the formation of membrane components. Molecules produced by the RER and SER can be packaged into vesicles and sent to other locations in the cell or to the outside of the cell.) and the progesterone receptor (PR).

HER2 is a growth factorA substance that stimulates cell division. Growth factors are usually small proteins or steroid hormones. They may be secreted by the same cells on which they act or by cells that reside in a different part of the body than the target cells. Some examples of growth factors include estrogen, a growth factor for breast cells, and VEGF, a growth factor that causes the development of blood vessels. Several different anti-cancer treatments are designed to inhibit the activity of growth factors. receptor located on the surface of breast cells. About 30% of breast cancer patients have tumors that express a very large amount of HER2. The anti-cancer drug Herceptin® is targeted at this protein making accurate determination of the presence or absence of this protein in a tumor an important step in determining the appropriate treatment.
Learn more about Herceptin®
Learn More About HER2

The ER protein is another growth factor receptor. The ER protein binds to the female sex hormoneA chemical produced by cells that alters the activity of other cells. The chemicals may be lipids, such as testosterone and estrogen or proteins like insulin. Hormones may act at locations far from their site of origin. Estrogen, for example, is produced primarily by cells in the ovaries but acts on cells in the breast and elsewhere. estrogen and plays a major role in stimulating cell division in breast cells. Because drugs that interfere with estrogen signaling are an important tool in treating breast cancer, accurate determination of ER levels is important to the design of treatment plans.
Learn more about hormonal treatments for breast cancer

The PR protein is the receptor for the female sex hormone progesterone. While there are no targeted therapies directed against the PR protein, the presence or absence of the receptor in cancer cells is a factor in determining the prognosis of the disease. Learn More About ER and PR

A Closer Look at Immunohistochemistry Results

Scoring system and assessment for HER2 status

• Scoring (from the makers of HercepTest™)
• (0) - one or little staining in < 10% of cells
• (1+) - faint, partial staining in > 10% of cells
• (2+) - weak to moderate, complete staining in > 10% of cells
• (3+) - strong, complete membrane staining in > 10% of cells
• Assessment of staining (according to the NCCN)
• Samples with 3+ (Not 2+/3+ borderline) are eligible for Herceptin™
• Samples with 2+ result should be retested using FISH
• Samples with 0/1+ result are HER2 negative

Scoring system and assessment for( )ER, PR status

• J-Score Method Scoring
• (0) - no stained cells
• (1+) - stained cells ≤ 1%
• (2+) - 1% < stained cells < 10%
• (3+) - stained cells ≥ 10%
• Assessment of staining
• Negative - Score of 0
• Indeterminate - Score of 1 or 2
• Positive - Score of 3

• Allred Method Staining
• (0) - no stained cells
• (1) - stained cells <1/100
• (2) - 1/100 ≤ stained cells < 1/10
• (3) - 1/10 ≤ stained cells < 1/3
• (4) - stained cells = 1/3 < 2/3
• (5) - stained cells > 2/3
• Intensity
• 0 = none
• 1 = weak
• 2 = intermediate
• 3 = strong
• Scoring
• To obtain the total score add staining score and intensity score. Any score between 0-2 is considered( )ER or PR negative, any score above 2 in considered( )ER or PR positive.