Immunohistochemistry stains

Author: Brian Wu PhD. MD Candidate, Keck School of Medicine; Chief Editor: Dr Amanda Oakley, Dermatologist, Hamilton, New Zealand, July 2015.

Immunohistochemistry stains — codes and concepts

What is immunohistochemistry?

Immunohistochemistry (IHC) is considered to be an advanced form of histopathology. It is not usually used initially, but rather added when routine/regular histological testing cannot provide the answers which clinicians need to form a diagnosis – and thus a plan of care.

IHC uses primary antibodies to label a protein, then follows this up with a secondary antibody which is bound to the primary one. In immunoperoxidase staining, an antibody is joined to a enzyme, peroxidase, that catalyses a reaction in which the protein is specifically stained brown. IHC can also involve fluorescently labelled antibody, so that when viewed under a light microscope, a certain pattern will be observed from the emitted fluorescence.

The IHC pattern is considered diagnostic. This is why IHC is often used in situations where a presence or absence of certain proteins can form a basis for a diagnosis. It can also be used to distinguish between two different disease processes that may otherwise appear similar to the pathologist.

How is an immunohistochemistry assay performed?

The most common process of preparing immunohistochemical slides is as follows:

  1. Fixation of the tissue (in general, IHC stains are fixed with formalin)
  2. Embedding of the tissue (in paraffin)
  3. Sectioning of the tissues
  4. Retrieval (done with application of heat or proteolytic enzymes)
  5. Mounting and dehydration
  6. Clearing and observation of the slides.

What are some examples of immunohistochemistry stains?

A wide variety of immunohistochemical stains is used to identify different tumours and other neoplasms. Some of the most common are listed in the table below.

IHC StainUses
Alpha-fetoprotein Used to detect hepatocellular carcinomas and germ cell neoplasms  
Alpha-1-antichymotrypsin Can identify soft tissue histiocytomas and fibroxanthomas of the skin  
BCL2 Used to distinguish between basal cell carcinomas and trichoepitheliomas
Image is of primary cutaneous diffuse large B cell lymphoma
Primary cutaneous diffuse large B cell lymphoma bcl 2 stain
Calponin Can differentiate between benign and malignant breast lesions  
CD30 Can be used in the diagnosis of Hodgkin lymphoma and anaplastic lymphomas
Image is of lymphomatoid papulosis
Lymphomatoid papulosis pathology CD30+ stain
CD31 Helps to identify endothelial tumors
Image is of epithelioid angiosarcoma
CD31 positive in epithelioid angiosarcoma
CD34 Distinguishes different endothelial tumours
Image is of the cellular variant of dermatofibroma
CD34+ in cellular variant of dermatofibroma
CD56 Used in the diagnosis of non-Hodgkin lymphomas, leukemias and small cell carcinomas  
Chromogranin Can mark some endocrine tumours  
EMA Used to identify eccrine neoplasms, Paget disease and sebaceous carcinomas
Image is of clear cell acanthoma
EMA+ in clear cell acanthoma
Factor 8 Used to mark megakaryocytes and endothelial cells  
Factor 13 Can help clinicians distinguish between dermatofibrosarcomas and dermatofibromas
Image is of dermatofibroma
Factor 13A+ in dermatofibroma
HMB 45 Used to detect melanocytes, naevus cells and melanomas
Image shows staining in melanoma and not in adjacent naevus
Melanoma stains HMB 45+, and adjacent naevus is negative
Cytokeratin High Molecular Weight Used to detect ductal carcinomas, squamous cell carcinomas and other epithelial neoplasms
Image is of Pagetoid infiltration of the epidermis by breast cancer
CK7+ in Pagetoid infiltration of epidermis by breast cancer
Cytokeratin 20 Can help identify adenocarcinomas of the gastrointestinal and reproductive system as well as gastrointestinal epithelial tumors
Image is of Merkel cell carcinoma
CD20+ in Merkel cell carcinoma pathology
MART, Melan-a Can help identify melanocytic naevus cells and melanomas  
S-100 Used to mark tumours of the melanocytes, both naevi and melanomas
Image is of folliculotropic metastatic melanoma
S100+ in folliculotropic metastatic melanoma
Vimentin Helps identify mesenchymal tumors  

What are the advantages and disadvantages of immunohistochemistry?

The advantages of IHC include:

  • It is possible to use fresh or frozen tissue samples for IHC.
  • IHC is well-established and readily available.
  • The cost of IHC is relatively low
  • It has a fast turn-around time.
  • Because there are no infectious agents involved in the study, the risk to human health is minimal.

The disadvantages of IHC are as follows:

  • IHC stains are not standardised worldwide.
  • While the cost of the procedure is relatively inexpensive, the equipment needed to perform IHC is costly.
  • Quantifying results is difficult.
  • IHC is subject to human error. Well-trained personnel are paramount.


Related information



  • Grkovic, I. Immunohistochemistry. Department of Anatomy, Histology and Embryology, University of Split. 2007
  • IHC and ISH advanced staining. Leica Biosystems. 2014
  • Immunohistochemical stains. Stanford University. 2013.
  • Kiupel, M. Diagnostic molecular pathology. Diagnostic Center for Population and Animal Health, College of Veterinary Medicine, Michigan State University, 2010.
  • Peckham, A. Immunohistochemistry. Faculty of Biological Sciences, Leeds University, 2003.
  • Special stains and immunohistochemistry. Central Coast Pathology Laboratory. 2015.

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