Chromogenic reporters, also known as chromogens, are chemical compounds used in immunohistochemistry (IHC) labeling procedures to visualize the presence of specific target molecules in tissue samples. These chromogenic reports work by undergoing a chemical reaction with enzymes linked to secondary antibodies, producing a visible color change or colored precipitate that can be visualized by light microscopy. The enzymes commonly used in chromogenic detection are horseradish peroxidase (HP) and alkaline phosphatase (AP).
The three most common chromogenic reporters used in immunohistochemistry (IHC) include:
- DAB (3,3'-diaminobenzidine):DAB produces an intense brown, permanent precipitate on reacting with horseradish peroxidase enzyme. Its longevity and resistance to fading on light exposure make it the preferred chromogenic reporter for IHC.
- AEC (3-amino-9-ethylcarbazole):On reacting with horseradish peroxidase enzyme, AEC produces an intense red precipitate that contrasts well with blue in double staining.
- BCIP/TNBP: BCIP/TNBP reacts with alkaline phosphatase enzymes to generate an intense blue color.
The choice of chromogenic reporter used is directly related to the type of test and the enzyme used in the test. Each enzyme is capable of catalyzing the conversion of specific substrates.