Immunohistochemistry for Pathologists: Protocols, Pitfalls, and Tips

Abstract

Immunohistochemistry (IHC) is an important auxiliary method for pathologists in routine diagnostic work as well as in basic and clinical research including exploration of biomarkers, as IHC allows confirmation of target molecule expressions in the context of microenvironment. Although there has been a considerable progress in automation and standardization of IHC, there are still many things to be considered in proper optimization and appropriate interpretation. In this review, we aim to provide possible pitfalls and useful tips for practicing pathologists and residents in pathology training. First, general procedure of IHC is summarized, followed by pitfalls and tips in each step and a summary of troubleshooting. Second, ways to an accurate interpretation of IHC are discussed, with introduction to general quantification and analysis methods. This review is not intended to provide complete information on IHC, but to be used as a basic reference for practice and publication.

Keywords: Antigen-antibody reactions; Auxiliary test; Immmunohistochemistry; Immunostain.

Fig. 1.

Illustration of polymeric amplification system. DAB, diaminobenzidine; HRP, horseradish peroxidase.

Fig. 2.

Receiver operating characteristic curve. Test A that is closer to the upper left-hand corner is better than test B and C. Performance of the test A is also better than that of the other two tests considering the area under the curve. Test D, lined in the opposite side of the reference line, represents negative correlation with the predetermined gold standard. TPR, true positive rate; FPR, false positive rate.