Reevaluation of assay methods and establishment of kit for superoxide dismutase activity.
Reevaluation of assay methods and establishment of kit for superoxide dismutase activity.
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DOI：
10.1016/0003-2697(84)90467-6
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发表时间：
1984-11
影响因子：
2.9
通讯作者：
Y. Ōyanagui
中科院分区：
文献类型：
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作者：
Y. Ōyanagui
超氧化物歧化酶 (SOD) 活性通过七种测定方法进行测量。我们发现亚硝酸盐法最适合我们的 SOD 检测试剂盒。然后对该方法进行了修改，以提供更好的灵敏度并最大限度地减少共存蛋白质的干扰，这是一个以前被忽略的因素。羟胺或其 O-磺酸、黄嘌呤氧化酶、次黄嘌呤、EDTA 和样品在有或没有 KCN 的情况下在 pH 8.2、37°C 下孵育 30 分钟。添加重氮染料形成试剂并在550 nm处测量吸光度。使用该 SOD 试剂盒和细胞色素 c 方法对健康成人和唐氏综合症患者的人血浆和红细胞裂解物进行测定。我们的试剂盒的灵敏度比细胞色素 c 方法高 8.5 倍。这种高灵敏度允许使用简单的分光光度计，而且只需一次稀释即可借助我们的公式确定 SOD 单位。证明了试剂具有良好的回收率、重现性和稳定性。
Superoxide dismutase (SOD) activity was measured by seven assay methods. The nitrite method was found to be the best for our SOD assay kit. This method was then modified to give better sensitivity and minimize interference by coexisting protein, a factor which has been previously ignored. Hydroxylamine or its O-sulfonic acid, xanthine oxidase, hypoxanthine, EDTA, and the sample were incubated with or without KCN at pH 8.2, 37°C, for 30 min. Diazo dye-forming reagent was added and the absorption was measured at 550 nm. Human plasma and erythrocyte lysate from healthy adults and Down's syndrome patients were assayed by this SOD kit and by the cytochrome c method. Our kit gave 8.5 times higher sensitivity than the cytochrome c method. This high sensitivity allowed the use of a simple spectrophotometer and, moreover, only one dilution was needed to determine the SOD unit with the help of our formulas. Good recovery, reproducibility, and stability of reagents were demonstrated.