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Luminol-enhanced Assay for Superoxide Anion (O2 face="Symbol" ,,,)


Non-cytotoxic chemiluminescent enhancer for increased sensitivity

Luminol-enhanced Assay for Superoxide Anion (O2 )

Danny Q. Hoang
Stratagene

Lorraine C. Pfefferkorn
Dartmouth School of Medicine, Dept. of Microbiology, Hanover, NH

Stratagene introduces the LumiMax detection assay, a simple and unique chemiluminescent-based method to qualitatively detect the presence of superoxide anions in cell culture. In addition, the kit provides a specific enhancement reagent to amplify chemiluminescent signal output that is noncytotoxic and nondenaturing to live cells. Superoxide anion (O2 ), a potent oxidant, is a short-lived oxygen radical that is released into the extracellular environment of stimulated leukocytes (monocytes, macrophages, and polymorphonuclear leukocytes).1 Superoxide anion is synthesized when NADPH-oxidase, a plasma membrane enzyme complex containing cytochrome b 558, transfers an electron to molecular oxygen. Superoxide anions are implicated in several key cellular processes, including oxidative stress damage,2 tumor promotion,3 and, most recently, cell growth and DNA synthesis via their involvement in the cell signaling pathway of proto-oncogene ras.4

At present, superoxide may be detected by measuring the reduction of exogenously supplied cytochrome c,5 the uptake of oxygen from the medium (measured by the Clark electrode6), or luminol-mediated chemiluminescence.7 The first method has limited sensitivity and is complicated by the reoxidation of superoxide-reduced cytochrome c by contaminants and cell lysis. The oxygen-uptake method is not only less sensitive than cytochrome c reduction
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