Unlike chemiluminscent or fluorescent blotting applications, chromogenic substrates do not require special equipment for visualization of the assay results. Similar to developing film, the blot is incubated in substate until the desired amount of development. Unlike chemiluminescent blotting assays, the colored precipitate cannot be easily be stripped off for reprobing.

Protein method: Chromogenic Western Blotting Substrates

Which chromogenic detection kit is right for you?

  1-Step Chloronaphthol Metal-enhanced DAB Substrate Kit CN/DAB Substrate Kit 1-Step Ultra TMB Blotting Solution
Format Ready-to-use liquid 2-component
reagent kit
reagent kit
Ready-to-use liquid
Features Stable at 4°C for 1 year Working solution is stable for over six hours at RT Stable at 4°C for 1 year Stable at 4°C for 1 year
Sensitivity 5 ng 17 pg 500 pg 20 pg
Color Blue-purple Brown-black Black Dark blue
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Additional chromogenic kits

WesternBreeze® Chromogenic Kits

WesternBreeze® Chromogenic Kits yield high-sensitivity results with extremely low background without the need for any additional equipment—just watch the signal develop over a short period of time. The kit uses the chromogenic phosphatase substrate BCIP (5-bromo-4-chloro-3-indolyl phosphate) and electron-transfer agent NBT (nitro blue tetrazolium) to produce a dark blue precipitate at the precise site of enzymatic activity on the blot.

The WesternBreeze® kit has been optimized to provide picogram sensitivity. Each kit includes ready-to-use or easy-to-dilute blockers, primary antibody diluent, wash solution, conjugated secondary antibody solution, substrate, and two convenient incubation trays.


Pierce™ NBT and BCIP Substrates are ready-made solutions of nitro-blue tetrazolium and 5-bromo-4-chloro-3'-indolyphosphate for chromogenic blot and IHC staining with alkaline phosphatase probes.

The combination of NBT (nitro-blue tetrazolium chloride) and BCIP (5-bromo-4-chloro-3'-indolyphosphate p-toluidine salt) yields an intense, insoluble black-purple precipitate when reacted with alkaline phosphatase, a popular enzyme conjugate for antibody probes. Pierce™ 1-Step NBT/BCIP solutions are available with and without a levamisole suppressor of endogenous phosphatase activity.