Assay Principle: Luminol reacts with superoxide and other reactive oxygen species resulting in a luminophore that has an emission peak at ~425 nm. The intensity of the chemiluminescence is proportional to the amount of superoxide or reactive oxygen species in the sample. The superoxide reaction shown above exemplifies chemiluminescence production from luminol.
The kit is based on a proven assay which is used to quantify gross cellular production of reactive oxygen species including; superoxide, hydroxyl radical, peroxynitrite and hypochlorite. When the other reactive oxygen species are mostly derived from superoxide, this assay provides an indirect measure of cellular superoxide production. Indeed, luminol assay is one of the most important methods in the analysis of metabolic activities of phagocytes and is widely used in biomedical research.

WPI Enhancer: A number of compounds are known to increase the intensity of luminescent emission from luminol. The WPI SuperLuminol kit utilizes a patented non-toxic enhancer to provide ~50 times increase in luminescence emission intensity from cells. The figure above shows oxidative bursts from phagocytes in whole blood (after 400 times dilution) induced by phorbol myristate acetate (PMA) in the absence and presence of the enhancer. Significantly lower amounts of reactive oxygen species can be assayed when using the enhancer.

Kit Contents: The WPI SuperLuminol kit contains a complete reagent set to assay superoxide/reactive oxygen species for cell cultures, isolated phagocytes, un-isolated phagocytes in whole blood, and other biological samples. The kit can be purchased with or without a chemical stimulant/agonist (PMA) for superoxide/reactive oxygen species production in cells. 

The kit includes the following reagents:

• Luminophoric Reagent: Luminol
• Assay Buffer: HBSS
• Enhancer
• KO2 (qualitative test control reagent)
• PMA (optional)
• PMA solvent (dimethyl-sulfoxide, optional)

Benefits:

• Simple and extensively tested method with an increased sensitivity than chemical methods
• Further improvement of sensitivity using a proprietary enhancer
• Minimal cellular toxicity from luminol and the enhancer

Caldefie-Chezet, et al. (2002) “Is the neutrophil reactive oxygen species production measured by luminol and lucigenin chemiluminescence intra or extracellular? Comparison with DCFH-DA flow cytometry and cytochrome c reduction”, Clin. Chim. Acta 319, 9-17.
Lojek, et al. (2002) “A comparison of whole blood neutrophil chemiluminescence measured with cuvette and microtitre plate luminometers”, Luminescence 17, 1-4.
Faulkner & Fridovich (1993) “Luminol and Lucigenin as detectors for O2- ”, Free Radic. Biol. Med. 15, 447-451.
Fridovich (1997) “Superoxide anion radical (O2&Mac183;-), superoxide dismutases and related matters”, J. Biol. Chem. 272, 18515-18517.