Immunohistochemistry (IHC) is a diagnostic technique that entails the detection of a target antigen (usually a protein) on tissue sections using antibodies. The tissue antigen is recognized by a specific antibody added to the tissue section. The antigen-antibody reaction is then visualized using a histochemical (enzyme-substrate-chromogen) reaction. The visualization of the antigen in a particular tissue or cell is the basis for diagnoses that are not possible with routine histopathology.

Uses of IHC in veterinary medicine:

1. Diagnosis of neoplasia. The use of IHC in neoplasms is based on the unique antigens expressed by different types of neoplastic cells and by their nonneoplastic cells of origin. Exploiting this feature, diagnosticians can classify leukocytic tumors (e.g., histiocytic sarcoma, lymphoma, mast cell tumor). Tumors that are morphologically similar (tumor mimics) but have a very different prognosis (e.g., epitheliotropic lymphoma and cutaneous histiocytoma in dogs) can be distinguished with immunohistochemistry. The improved specificity in the diagnosis of neoplasia with IHC can improve cancer treatment. Unfortunately, the use of only one IHC test is not helpful in many cases due to “antigen sharing” among different cell types/tumors. Therefore, a panel of IHC tests is typically required for definitive diagnosis.

2. Diagnosis of carcinomas of unknown primary site (CUPS). One of the main challenges a diagnostician faces is the evaluation of tumors simultaneously present in several organs, making determination of the primary site difficult with routine histopathology, particularly with biopsy samples. IHC may aid in the determination of the primary site.

3. Prognosis and theranostics¹. Although the use of IHC for these purposes is well established for many human tumors, advances have also been made in small animal oncology. Examples are prognostic markers used for canine melanoma, mast cell tumor, osteosarcoma, and mammary tumors. Proliferation index is the most common prognostic marker used in a wide variety of animal cancers. An example of biomarkers used in animal theranostics is KIT. This antigen has variable intracellular expression depending on the mast cell tumor grade, which is used to customize therapy for a given tumor.

4. Diagnosis of infectious diseases. IHC can identify microorganisms (e.g., virus, bacteria, protozoa, prions) in tissue sections. Although the use of IHC in infectious diseases has been largely replaced by more sensitive and specific techniques (e.g., PCR), the ADDL offers IHC tests for some microorganisms. IHC is less sensitive than PCR, but allows the co-localization of microorganisms in specific cell types or within a lesion, which is not possible with routine PCR methodologies.

Interpretation of results:

IHC results are part of the final diagnosis and only exceptionally considered a stand-alone technique. Therefore, a pathologist must evaluate not only the IHC sections but also the routine histopathology and any other ancillary tests in a given case. In other words, IHC is complementary to the other techniques available to a pathologist.

Immunohistochemistry at ADDL:

IHC has been offered at ADDL for more than 10 years. To search for available tests, visit (http://www.addl.purdue.edu/TestsFees/BySection.aspx)  In most cases, IHC is performed after the pathologist reviews routine histopathology sections. IHC tests are not included in the standard pathology (necropsy, mail-in, biopsy) fee, so clients are contacted for permission to run these additional tests. Samples for IHC are processed in the same way as for routine histopathology so there are no special submission procedures other than prompt immersion of the sample in an adequate volume of formalin fixative.

The ADDL has recently purchased a new IHC stainer that has improved the turnaround time of results. Keep in mind that the list of available IHC tests may change over time depending on our clients’ needs. Please contact the ADDL to confirm that the requested test is available. Your feedback is very much appreciated. Email us at addl@purdue.edu.