Western Blot - 5. Chemiluminescent substrates:

Rockland produces several luminol based Chemiluminescent substrate for the detection of horseradish peroxidase (HRP).  PicoMax™ and FemtoMax™ are designed for high performance in western blotting and is functional on both nitrocellulose and PVDF membranes. FemtoMax™ produces chemiluminescence and allows for the detection of down to femtogram (10-15) amounts of antigen. Detection methods may include photographic film or other imaging methods, including highly sensitive CCD camera based systems.

Chromogenic substrates:

Chromogenic blotting substrates are available from Rockland in a variety of specifications and formats. The appropriate substrate choice depends on the enzyme label, desired sensitivity and form of signal or method of detection needed.   

Chromogenic Peroxidase Substrates:

The Peroxidase reaction with our TMBM substrate produces a water-soluble blue product that can be precipitated onto a membrane.  The precipitating product produces blue to dark blue bands in the enzyme location.  TMBM is well suited to applications that require high signal-to-noise. DAB is another peroxidase substrate and yields a brown precipitate in the presence of HRP and peroxide.  



Chromogenic Alkaline Phosphatase Substrates:  

The NBT/BCIP reagent is also commonly used in chromogenic western blot assays.  NBT serves as an oxidant and BCIP as the alkaline phosphatase substrate.  Together NBT and BCIP form reactants in the presence of alkaline phosphatase which yields a dark purple to black, water-insoluble, precipitant product providing strong sensitivity. 

Western Blot Protocol

1. Background
2. Types of Western Blot Formats
3. Primary Antibody Reagents
4. Secondary Reporter Antibody Reagents
5. Substrates
6. Buffers and Blocking Reagents
7. Western Blot Preassembled Kits