5-Bromo-4-chloro-3-indolyl phosphate disodium salt (BCIP disodium salt) is a widely used chromogenic substrate for the detection of alkaline phosphatase (AP) activity in molecular biology, immunohistochemistry, and diagnostic applications.
Upon enzymatic dephosphorylation by alkaline phosphatase, BCIP produces an indoxyl intermediate that undergoes oxidation and dimerization, forming an insoluble blue/purple precipitate. When used in combination with NBT (Nitro Blue Tetrazolium), signal intensity is significantly enhanced, producing a strong and stable colorimetric endpoint.
BCIP disodium salt is water-soluble and ideal for membrane-based assays, histological staining, and immunoassays requiring visible detection.
BCIP is a chromogenic substrate for alkaline phosphatase. Under the catalysis of alkaline phosphatase, BCIP undergoes hydrolysis:
| Parameter | Recommended Condition |
| Detection Wavelength | 595–620 nm (visible blue/purple precipitate) |
| Buffer System | Tris-HCl 0.1 M, pH 9.0–9.5; avoid phosphate buffers |
| Temperature | 20–25°C (room temp) or 30–37°C for faster kinetics |
| Assay Format | Membrane staining, tissue IHC/ISH, microplate colorimetric assay |
✅ Notes:
Visible colorimetric detection of alkaline phosphatase activity
No need for fluorescent or specialized imaging equipment
Cost-effective substrate for AP-based assays
Well-established in molecular biology and immunoassay literature
Storage conditions: Store powder at -20°C, protected from light and moisture
Prepare working solutions fresh or as instructed in the manual
Avoid repeated freeze-thaw cycles
Protect from light to prevent product fading
