Chromogens in Multiple Immunohistochemical Staining Used for Visual Assessment and Spectral Imaging: The Colorful Future

Abstract

For the chromogenic visualization of immunohistochemical enzymatic reaction products, only a limited series of different enzymatic activities and chromogens are available. Consequently, combinations of two chromogens for double immunohistochemical staining are even more limited for visual assessment. The recent introduction of spectral imaging allows unmixing of multiple immunohistochemically stained tissue samples for the exclusive observation of colocalization and quantitative measurement of individual staining components. Because unmixing is based on spectral characteristics, multiple immunohistochemistry is no longer dependent on chromogens showing a high visual contrast. Although this allows greater flexibility for combining chromogens in multiple staining applications, it also appears that some chromogens do not have excellent spectral and microscopic properties. The present work provides an overview of currently applied chromogens for single and multiple immunohistochemistry. (The J Histotechnol 33(1):31–40, 2010)

Submitted November 30, 2009; accepted January 30, 2010.

Keywords:

• chromogen
• immunohistochemistry
• multiple staining
• spectral imaging