What is Immunohistochemistry?
Immunohistochemistry (IHC) is a laboratory technique used in histology to detect specific antigens in tissue sections by using antibodies tagged with a visible label. It combines anatomical, immunological, and biochemical techniques to visualize the distribution and localization of specific cellular components within a tissue.
Why is Immunohistochemistry Important?
IHC is crucial for diagnosing diseases, particularly in
pathology. It helps in identifying abnormal cells in diseases such as cancer, and it is also used for research purposes to understand the spatial distribution of specific proteins, enzymes, and other molecules within tissues.
How Does Immunohistochemistry Work?
The basic principle of IHC involves the binding of an antibody to a specific antigen in the tissue. These antibodies can be
monoclonal or
polyclonal. The antibody is usually linked to a detectable marker such as an enzyme (e.g., horseradish peroxidase) or a fluorescent dye. When the antibody binds to the antigen, the marker can be visualized under a microscope, usually with the aid of a substrate that produces a colored reaction.
Fixation: Preserving the tissue to maintain cellular structure and antigenicity.
Embedding: Embedding the fixed tissue in paraffin wax to facilitate sectioning.
Sectioning: Cutting thin slices of the tissue for mounting on slides.
Deparaffinization and Rehydration: Removing the paraffin and rehydrating the tissue sections.
Antigen Retrieval: Unmasking antigens to enhance antibody binding, often through heat-induced methods.
Blocking: Preventing non-specific binding of antibodies.
Primary Antibody Incubation: Applying the primary antibody that binds to the antigen of interest.
Secondary Antibody Incubation: Applying a secondary antibody that binds to the primary antibody and is linked to a detectable marker.
Detection: Visualizing the antigen-antibody complex using appropriate substrates or fluorescent dyes.
Cancer Diagnosis: Identifying specific cancer markers to aid in diagnosis and treatment planning.
Research: Studying the expression and localization of proteins in various tissues.
Neuropathology: Investigating protein accumulations in neurodegenerative diseases.
Drug Development: Evaluating the effects of new drugs on targeted tissues.
Antibody Specificity: Non-specific binding can lead to false positives.
Antigen Retrieval: Some antigens may be difficult to unmask without damaging the tissue.
Quantification: IHC is generally qualitative, and quantifying protein levels can be challenging.
What are Recent Advances in Immunohistochemistry?
Recent advances have improved the sensitivity and specificity of IHC. Innovations include the development of
multiplex IHC techniques that allow the simultaneous detection of multiple antigens in a single tissue section, and the use of
digital pathology and image analysis software for more accurate quantification and interpretation of IHC results.
