2,7-Dihydrodichlorofluorescein diacetate as a fluorescent marker for peroxynitrite formation
2,7-Dihydrodichlorofluorescein diacetate as a fluorescent marker for peroxynitrite formation
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DOI：
10.1016/s0014-5793(97)01197-6
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发表时间：
1997-10-20
期刊：
影响因子：
3.5
通讯作者：
Wolf, G
中科院分区：
文献类型：
--
作者：
Possel, H;Noack, H;Wolf, G
活性氧 (ROS) 被认为是细胞损伤（包括细胞凋亡和坏死）的重要致病因素，它们的作用包括脂质过氧化、DNA 损伤、线粒体呼吸链破坏和蛋白质修饰。最近的实验强调了过氧亚硝酸盐的重要性，过氧亚硝酸盐是两种有效活性物质一氧化氮和超氧化物的反应产物，几种 荧光化合物已被用于测定活细胞中ROS的形成，除了目前主要用于监测过氧亚硝酸盐的二氢罗丹明-123（DHR-123）外，2,7-二氢二氯荧光素（DCF-H）用于检测过氧化氢和一氧化氮，我们采用无细胞方法来评估其特异性和灵敏度 DCF-H 对各种氧化化合物，我们的研究表明 DCF-H 对过氧亚硝酸盐氧化比任何其他测试的化合物更敏感。为了研究单个细胞内过氧亚硝酸盐的生成，仅用激光扫描显微镜监测负载 DCF-H 的原代胶质细胞培养物，刺激小胶质细胞同时产生过氧亚硝酸盐前体一氧化氮和超氧化物，显示 DCF 荧光的最大增加，而单独产生一氧化氮或超氧化物的小胶质细胞显示相对较小的增加 在 DCF 荧光中。结论是，DCF-H 被证明是一种优秀的过亚硝酸盐标记物，具有检测活细胞中过亚硝酸盐形成的潜力。 (C) 1997 年欧洲生化学会联合会。
Reactive oxygen species (ROS) have been implicated as an important causative factor in cell damage, including apoptosis and necrosis, Their proposed actions comprise lipid peroxidation, DNA damage, destruction of the mitochondrial respiratory chain and protein modifications, Recent experiments underline the importance of peroxynitrite, the reaction product of the two potent reactive species nitric oxide and superoxide, Several fluorogenic compounds have been used in order to determine ROS formation in living cells, Besides dihydrorhodamine-123 (DHR-123), at present mostly applied to monitor peroxynitrite, 2,7-dihydrodichlorofluorescein (DCF-H) is used for detection of hydrogen peroxide and nitric oxide, We employed a cell free approach to evaluate the specificity and sensitivity of DCF-H to various oxidizing compounds, Our studies imply that DCF-H is much more sensitive to peroxynitrite oxidation than any other compound tested. In order to study peroxynitrite generation within individual cells, primary glial cultures loaded with DCF-H mere monitored with a laser scanning microscope, Microglia, stimulated to simultaneously produce the peroxynitrite precursors nitric oxide and superoxide, displayed the greatest increase in DCF fluorescence, whereas microglia producing either nitric oxide or superoxide alone showed a relatively small increase in DCF fluorescence. Tn conclusion, DCF-H was demonstrated to be an excellent peroxynitrite marker with the potential to detect peroxynitrite formation in living cells. (C) 1997 Federation of European Biochemical Societies.