Quantifying ROS levels using CM-H2DCFDA and HyPer
Quantifying ROS levels using CM-H2DCFDA and HyPer
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DOI：
10.1016/j.ymeth.2016.06.008
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发表时间：
2016-10-15
期刊：
影响因子：
4.8
通讯作者：
Wieckowski, Mariusz R.
中科院分区：
文献类型：
--
作者：
Oparka, Monika;Walczak, Jaroslaw;Wieckowski, Mariusz R.
在低水平下，活性氧簇(ROS)可以作为细胞内的信号分子。当ROS的产生大大超过内源抗氧化剂系统的能力，或者抗氧化剂水平降低时，ROS水平进一步增加。后者与氧化应激诱导和相关的信号转导有关，其特征是ROS诱导的细胞氧化还原动态平衡的变化和/或对生物分子(如DNA、蛋白质和脂类)的破坏作用。由于ROS的产生和去除涉及复杂的机制，再加上缺乏真正针对特定类型ROS的报告分子，量化(亚)细胞ROS水平是一项具有挑战性的任务。在本章中，我们描述了两种测量ROS的方法：一种方法使用化学报告CM-H(2)DCFDA(5-(and-6)-chloromethyl-2‘，7’-dichlorodihydrofluorescein(双乙酸酯)来评估一般氧化剂水平，另一种方法允许使用基于蛋白质的传感器(HYPER和SYPHER)更具体地分析胞液过氧化氢(H2O2)水平。(C)爱思唯尔公司出版的2016年。
At low levels, reactive oxygen species (ROS) can act as signaling molecules within cells. When ROS production greatly exceeds the capacity of endogenous antioxidant systems, or antioxidant levels are reduced, ROS levels increase further. The latter is associated with induction of oxidative stress and associated signal transduction and characterized by ROS-induced changes in cellular redox homeostasis and/or damaging effects on biomolecules (e.g. DNA, proteins and lipids). Given the complex mechanisms involved in ROS production and removal, in combination with the lack of reporter molecules that are truly specific for a particular type of ROS, quantification of (sub)cellular ROS levels is a challenging task. In this chapter we describe two strategies to measure ROS: one approach to assess general oxidant levels using the chemical reporter CM-H(2)DCFDA (5-(and-6)-chloromethyl-2',7'-dichlorodihydrofluorescein diacetate), and a second approach allowing more specific analysis of cytosolic hydrogen peroxide (H2O2) levels using protein-based sensors (HyPer and SypHer). (C) 2016 Published by Elsevier Inc.