基于流式细胞术的低氧条件下线粒体 ROS 产生检测方案。
A flow-cytometry-based protocol for detection of mitochondrial ROS production under hypoxia.
机构信息
State Key Laboratory of Biotherapy and Cancer Center, West China Hospital, Sichuan University and Collaborative Innovation Center of Biotherapy, NO.17, South of Renmin Road, Chengdu 610041, China.
Experimental and Research Animal Institute, Sichuan University, Chengdu, China.
出版信息
STAR Protoc. 2021 Apr 20;2(2):100466. doi: 10.1016/j.xpro.2021.100466. eCollection 2021 Jun 18.
Hypoxia is known to stimulate mitochondrial reactive oxygen species (mROS) in cells. Here, we present a detailed protocol to detect mROS using MitoSOX staining in live cells under normoxia and hypoxia. Flow cytometry allows sensitive and reliable quantification of mROS by FlowJo software. We optimized several aspects of the procedure including hypoxic treatment, working concentrations of the staining buffer, and quantitative analyses. Here, we use HepG2 cells, but the protocol can be applied to other cell lines. For complete details on the use and execution of this protocol, please refer to Yang et al. (2020).
缺氧已知会刺激细胞中线粒体活性氧 (mROS)。在这里，我们介绍了一种使用 MitoSOX 染色在常氧和缺氧条件下活细胞中检测 mROS 的详细方案。流式细胞术允许通过 FlowJo 软件对 mROS 进行灵敏和可靠的定量分析。我们优化了该程序的几个方面，包括低氧处理、染色缓冲液的工作浓度和定量分析。在这里，我们使用 HepG2 细胞，但该方案可应用于其他细胞系。有关该方案使用和执行的完整详细信息，请参阅 Yang 等人。（2020 年）。
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