Kato Takamitsu A
Department of Environmental and Radiological Health Sciences, Colorado State University, Fort Collins, CO, USA.
Methods Mol Biol. 2025;2933:87-91. doi: 10.1007/978-1-0716-4574-1_12.
Cells manage oxidative stress from endogenous and exogenous stimuli. Excess oxidative stress causes significant cellular health issues, impacting key biological components such as DNA, membrane, and proteins. Reactive oxygen species (ROS) are a major contributor to oxidative stress, making the measurement of oxidative stress crucial for monitoring cellular health. DCFDA (2',7'-dichlorofluorescin diacetate) is a widely used probe for detecting oxidative stress in live cells. Once inside the cell, DCFDA is converted to DCFH, which is then oxidized to produce the fluorescent compound DCF. The green fluorescence emitted by DCF under blue light exposure serves as an indicator of oxidative stress. However, DCFDA is a broad oxidative probe and does not provide specificity for different types of ROS. This chapter introduces the use of carboxy-HDCFDA, a derivative of DCFDA, for monitoring HO-induced oxidative stress. Using a fluorometer, the fluorescence of carboxy-HDCFDA can be measured to provide insights into oxidative stress levels in cells. This approach allows for more precise monitoring and analysis of oxidative stress, enhancing our understanding of cellular responses to oxidative damage.
细胞应对来自内源性和外源性刺激的氧化应激。过量的氧化应激会引发严重的细胞健康问题，影响诸如DNA、细胞膜和蛋白质等关键生物成分。活性氧（ROS）是氧化应激的主要促成因素，因此氧化应激的测量对于监测细胞健康至关重要。2',7'-二氯荧光素二乙酸酯（DCFDA）是一种广泛用于检测活细胞中氧化应激的探针。一旦进入细胞，DCFDA会转化为DCFH，然后被氧化生成荧光化合物DCF。在蓝光照射下DCF发出的绿色荧光可作为氧化应激的指标。然而，DCFDA是一种宽泛的氧化探针，对不同类型的ROS没有特异性。本章介绍了使用DCFDA的衍生物羧基-HDCFDA来监测HO诱导的氧化应激。使用荧光计可以测量羧基-HDCFDA的荧光，以深入了解细胞中的氧化应激水平。这种方法能够更精确地监测和分析氧化应激，增进我们对细胞对氧化损伤反应的理解。
