Alkaline Phosphatase based immunostaining systems are gaining wide acceptance among pathologists because of their relatively high signal to noise ratio. 5-bromo,4-chloro,3-indolylphosphate BCIP)/iodoblue tetrazolium (INT) substrate for alkaline phosphatase-based detection systems, is a commonly used substrate chromogen. BCIP acts as the substrate for alkaline phosphatase, and the INT enhances the yellow-brown final color of the precipitates at the reaction sites. This chromogen is useful for immunohistochemistry, immunocyto-chemistry and in situ hybridization procedures in both manual and automated systems.
Slides should be counterstained with Biømeda Aqueous Hematoxylin (Cat. Np. M10) and mounted
in Crystal/Mount (Cat. No. M02). BCIP/INT is not compatible with organic solvents.
Refrigerate at 4°C. Do not freeze.
Using the Chromogen-Substrate:
Remove the BCIP/INT Chromogen-Substrate from the refrigerator 20 minutes before application.
1. Apply the Ready-to-Use BCIP/INT Chromogen Substrate to the specimen. Before applying this solution make sure to wash the specimens extensively with buffer to remove tracing reagent containing alkaline phosphatase.
2. Incubate for 15-30 minutes at room temperature. Wash specimens with distilled water and proceed with the counterstaining steps.