To determine SOD activity, several direct and
indirect methods have been developed. A common and convenient indirect
method utilizes nitroblue tetrazolium (NBT) conversion to NBT-diformazan
(formazan dye) via superoxide radical. However, there are several
disadvantages to the NBT method, such as poor water solubility of the
formazan dye and the interaction with the reduced form of xanthine
oxidase. Though cytochrome C is also commonly
used for SOD activity detection, its reactivity with superoxide is too
high to determine low levels of SOD activity. Cell Technology’s
SOD kit utilizes a water-soluble tetrazolium salt, WST-1
(2-(4-Iodophenyl)- 3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-
2H-tetrazolium, monosodium salt) that produces a highly water-soluble
formazan dye upon reduction with a superoxide anion (6). The rate of the
reduction with O2.- is linearly related to the
xanthine oxidase (XO) activity, and is inhibited by SOD, as shown in
Figure 1. Therefore, the IC 50 (50% inhibition activity of SOD or
SOD-like materials) can be determined by this colorimetric method.
Absorbance can be measured at 440nm.
Figure 1 - Inhibition Curve Prepared Using
SOD from Bovine Liver
Figure 2 - SOD ASSAY Reaction
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