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Molecular Labeling & Detection

NBT

Product information

NBT
Formula: C₄₀H₃₀NO₄BrClP x C₇H₉N
Molecular weight: 433.6

Applications

• Detection of alkaline phosphatase activity in:
  – Southern blotting,
  – Western blotting,
  – Northern blotting,
  – dot/slot blots,
  – plaque and colony screening.

Description

NBT (nitro blue tetrazolium) is an oxidant used in conjunction with BCIP or BCIP derivatives (BCIP T) (see protocol), to enhance blue color development during detection of alkaline phosphatase activity. It is suitable for immunohistochemistry and cytochemistry. NBT is also a substrate for oxidases and dehydrogenases.

Quality Control

One spot by TLC. Less than 8% water content.

Note

Preparation of a 75 mg/ml stock solution in 70% dimethylformamide is recommended.

Storage

Store at 4°C or at -20°C in the dark.

Patents, Licenses, Trademarks

Protocols & recommendations

RECOMMENDATIONS FOR USE

ADDITIONAL PROTOCOLS

Protocol for Preparation of BCIP-T/NBT Substrate Solution

Add the following components to 10 ml of the Alkaline Phosphatase (AP) buffer (100 mM Tris-HCl (pH 9.5), 100 mM NaCl and 10 mM MgCl₂):

BCIP-T (50 mg/ml in dimethylformamide)	33 µl
NBT (75 mg/ml in 70%dimethylformamide)	44 µl

Note

• Avoid exposure to light.
• Prepare fresh developing solution and use within an hour.
• BCIP-T and NBT solutions in dimethylformamide do not freeze. They are stable for approximately 2 years when stored at -20°C in the dark.