- 3-Amino-9-ethylcarbazole (AEC) is a chromogenic substrate commonly used in immunohistochemistry (IHC) and immunocytochemistry (ICC) for the detection of antigens when horseradish peroxidase (HRP) is employed as the enzyme label.
- In the presence of hydrogen peroxide (H₂O₂), HRP catalyzes the oxidation of AEC, producing a bright red, insoluble precipitate at the site of the antigen–antibody complex. This colored deposit marks the precise location of the antigen within the tissue or cell sample and can be visualized by light microscopy.
- One of the defining features of AEC is its color profile. Unlike 3,3′-diaminobenzidine (DAB), which produces a brown reaction product, AEC generates a vivid red stain that offers excellent contrast against blue nuclear counterstains such as hematoxylin. This makes it especially useful for highlighting antigens in tissues where a brown stain may be difficult to distinguish, or in cases where dual-staining approaches require distinct chromogens. The red coloration also makes AEC suitable for digital imaging and documentation in certain applications.
- AEC has some important limitations compared to DAB. The red precipitate produced is soluble in alcohol and organic solvents, which means that slides cannot be dehydrated or permanently mounted in non-aqueous media. Instead, aqueous mounting media must be used, and the staining is less stable over time, often fading during long-term storage. For this reason, AEC is typically used when immediate or short-term evaluation is required, rather than for archival purposes. Additionally, its sensitivity is somewhat lower than that of DAB, requiring careful optimization to achieve strong signal intensity.
- Despite these drawbacks, AEC is valued for specific applications in diagnostic and research pathology. Its strong contrast and distinct color make it particularly helpful in multiple-labeling experiments, where different chromogens are needed to detect more than one antigen in the same tissue section. It is also preferred in cases where permanent slide storage is not essential, or where its red signal provides better clarity for interpretation.
