Enzymes such as HRP or AP convert several substrates to a colored precipitate. As the precipitate accumulates on the blot, a colored signal develops. The reaction can be monitored and stopped when the desired signal over background is observed. Colorimetric detection is easier to perform than film-based chemiluminescence detection; however, the method's single endpoint result does not allow the multiple exposures of chemiluminescence methods. Colorimetric detection is typically considered a medium-sensitivity method compared to radioactive or chemiluminescence detection.
Bio-Rad's HRP-based colorimetric detection systems offer a choice of HRP-conjugated secondary antibodies and HRP-conjugated protein A or protein G using the detection reagent 4-chloro-1-naphthol(4CN). Bio-Rad offers three types of kits based on this chemistry for colorimetric HRP detection; individual reagents are also available, including 3,3'-diaminobenzidine (DAB), an alternative reagent.
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