Diagnostic Immunohistochemistry: What Can Go Wrong and How to Prevent It

Context: -There are a number of critical factors that can lead to incorrect results if the diagnostic pathologist performing immunohistochemistry is unaware of, or not vigilant about, their influence.

Objective: -To highlight 3 arenas in which errors may be introduced.

Data Sources: -For choosing the correct primary antibody, selection of the most appropriate antibodies for a given clinical application can be aided by obtaining information from the vendor; however, this can yield incomplete information. There are a number of online databases that have comparisons of antibodies from different vendors, particularly with respect to their use and properties. Reading the published literature can assist in this process, particularly with respect to determining antibody sensitivity and specificity, but it is a daunting task to keep up with all of the immunohistochemistry-related papers published. Finally, Web sites of a number of quality assurance organizations are accessible and can provide a wealth of information comparing the "real world" performance characteristics of different antibodies to the same target protein. False-positive signals can result from a number of factors, including the use of inappropriately high antibody concentration, and "pseudospecific" signal that is in the wrong compartment of the cell. False-negative signal can result from factors such as use of a nonoptimized epitope retrieval method. It is critical that epitope retrieval methods be optimized for each antibody employed in the laboratory.

Conclusions: -By paying attention to these potential problems, the "black box" of diagnostic immunohistochemistry can be made more transparent.