A flow-cytometry-based protocol for detection of mitochondrial ROS production under hypoxia

• Main Authors: Yun Yang, Guimin Zhang, Tao Yang, Jia Gan, Lin Xu, Hanshuo Yang
• Format: Article
• Language: English
• Published: Elsevier 2021-06-01
• Series: STAR Protocols
• Subjects: Cell Biology; Flow Cytometry/Mass Cytometry; Cell-based Assays; Molecular/Chemical Probes
• Online Access: http://www.sciencedirect.com/science/article/pii/S2666166721001738
• ISSN: 2666-1667

Summary: 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).