What factors should I consider when choosing enzyme and chromogen for the Ag-Ab reaction?

One factor is to ensure that the enzyme chosen is compatible with the substrate of the chromogen of interest. Different enzymes have specific substrates that produce distinct chromogenic reactions. For example, HRP is used for the DAB chromogen, while alkaline phosphatase is used for BCIP/NBT. Some enzymes may provide higher sensitivity and signal amplification compared to others. One should also assess the color intensity produced by the enzyme-chromogen reaction to provide a vibrant color such as Fast Red. Some enzymes may exhibit higher background staining compared to others, which can affect the specificity of the resulting staining.