The selective detection of mitochondrial superoxide by live cell imaging

被引：204

作者：

Robinson, Kristine M. ^{[1
]}

Janes, Michael S. ^{[1
,2
]}

Beckman, Joseph S. ^{[1
]}

机构：

[1] Oregon State Univ, Dept Biochem & Biophys, Linus Pauling Inst, Environm Hlth Sci Ctr, Corvallis, OR 97331 USA

[2] Invitrogen Mol Probes Labeling & Detect Technol, Eugene, OR 97402 USA

来源：

NATURE PROTOCOLS | 2008年 / 3卷 / 06期

关键词：

D O I：

10.1038/nprot.2008.56

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

A general protocol is described to improve the specificity for imaging superoxide formation in live cells via fluorescence microscopy with either hydroethidine (HE) or its mitochondrially targeted derivative Mito-HE (MitoSOX Red). Two different excitation wavelengths are used to distinguish the superoxide-dependent hydroxylation of Mito-HE (385-405 nm) from the nonspecific formation of ethidium (480-520 nm). Furthermore, the dual wavelength imaging in live cells can be combined with immunocolocalization, which allows superoxide formation to be compared simultaneously in cocultures of two types of genetically manipulated cells in the same microscopic field. The combination of these approaches can greatly improve the specificity for imaging superoxide formation in cultured cells and tissues.

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收藏

页码：941 / 947

页数：7

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