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Journal of Histochemistry and Cytochemistry, Vol. 45, 1665-1672, Copyright © 1997 by The Histochemical Society, Inc.

ARTICLE

Constant Detection of CD2, CD3, CD4, and CD5 in Fixed and Paraffin-embedded Tissue Using the Peroxidase-mediated Deposition of Biotin-Tyramide

Rainer Malisiusa, Hartmut Merza, Boris Heinza, Evariste Gafumbegetea, Britta U. Kocha, and Alfred C. Fellera
a Department of Pathology, Medical University of Lübeck, Lübeck, Germany

Correspondence to: Rainer Malisius, Dept. of Pathology, Medical University of Lübeck, Ratzeburger Allee 160, D 23538 Lübeck, Germany.

Immunohistochemical methods are widely used for diagnostic purposes in histopathology. However, the use of most monoclonal anti-leukocyte antibodies is limited to frozen tissues. Initially, it was believed that formalin fixation in particular, which is the gold standard for morphological tissue preservation, destroys most of the antigen binding sites. In recent years, protease digestion and the introduction of microwave techniques have significantly enhanced the sensitivity of immunohistochemical techniques, and a variety of hidden antigen sites in formalin-fixed tissue have been retrieved for initially unreactive antibodies. It therefore became clear that many of the leukocyte antigens are not irreversibly destroyed but are most probably masked during the fixation process. We developed a technique combining optimized pretreatment of formalin-fixed tissue with a dramatic enhancement of the immunohistochemical sensitivity and named it the ImmunoMax method. The ImmunoMax method proves that by optimizing the technique at the following three levels it is possible to detect formalin-sensitive leukocyte antigens: (a) standard fixation of the tissue; (b) sufficient antigen unmasking; and (c) increasing the substrate turnover by multiplication of binding sites with subsequent enhancement of the immunohistochemical reaction. Using this optimized ImmunoMax method, we were able to detect CD2, CD3, CD4, and CD5 with conventional monoclonal antibodies in formalin-fixed, paraffin-embedded tissue specimens of various lymphoid tissues. (J Histochem Cytochem 45:1665-1672, 1997)

Key Words: immunohistochemistry, biotin-tyramide, ImmunoMax, paraffin, CD2, CD3, CD4, CD5


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