Antigen retrieval

Antigen Retrieval is a technique in Immunohistochemistry used to unmask antigens in formalin-fixed, paraffin-embedded tissue sections, thereby enhancing the immunoreactivity of certain antibodies to their target antigens. This method is often necessary when the antigen of interest is not readily accessible due to the cross-linking of proteins and nucleic acids caused by formalin fixation.

History

The concept of antigen retrieval was first introduced in the early 1990s by Shi et al. Prior to this, the use of formalin-fixed, paraffin-embedded tissues in immunohistochemistry was limited due to the masking of antigenic sites.

Methods

There are two main methods of antigen retrieval: heat-induced antigen retrieval (HIAR) and proteolytic-induced antigen retrieval (PIAR).

Heat-Induced Antigen Retrieval (HIAR)

HIAR involves heating the tissue sections in a solution, often a citrate buffer. This process is thought to reverse the protein cross-linking caused by formalin fixation, thereby exposing the antigenic sites. HIAR can be performed using a microwave, pressure cooker, or water bath.

Proteolytic-Induced Antigen Retrieval (PIAR)

PIAR involves the use of proteolytic enzymes such as trypsin, pepsin, or proteinase K to digest the proteins and expose the antigenic sites. This method is less commonly used than HIAR due to the potential for over-digestion of the tissue.

Factors Influencing Antigen Retrieval

Several factors can influence the effectiveness of antigen retrieval, including the choice of retrieval solution, the temperature and duration of heating (for HIAR), and the concentration and incubation time of the proteolytic enzyme (for PIAR).

Applications

Antigen retrieval is widely used in diagnostic and research laboratories for immunohistochemical staining of formalin-fixed, paraffin-embedded tissues. It has greatly expanded the range of antibodies that can be used for immunohistochemistry, thereby enhancing our understanding of disease processes at the molecular level.

Immunohistochemical staining of a mouse brain section, demonstrating the use of antigen retrieval to enhance antibody binding.

See Also

• Immunohistochemistry
• Formalin-fixed, paraffin-embedded tissue
• Antibody

References

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