Detection of Intracellular Reactive Oxidative Species Using the Fluorescent Probe Hydroxyphenyl Fluorescein
Detection of Intracellular Reactive Oxidative Species Using the Fluorescent Probe Hydroxyphenyl Fluorescein
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DOI：
10.1007/978-1-0716-1258-3_18
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发表时间：
2021-01-01
期刊：
影响因子：
--
通讯作者：
Kawauchi，Keiko
中科院分区：
文献类型：
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作者：
Sugimoto，Wataru;Miyoshi，Daisuke;Kawauchi，Keiko
由于 ROS 水平与细胞状态密切相关，因此已开发出用于检测细胞内活性氧化物质 (ROS) 的各种荧光探针。在这里，我们描述了一种使用荧光探针羟苯基荧光素 (HPF) 检测活细胞中细胞内 ROS 的方法，该探针可检测羟基自由基和过氧亚硝酸盐。 NIH3T3 细胞和 p53 敲除 (p53−/−) 小鼠胚胎成纤维细胞 (MEF) 通过使用逆转录病毒系统表达致癌 RAS 进行转化。将细胞在 37°C 下用 HPF 处理 30 分钟，随后用 PBS 清洗后，使用共焦荧光显微镜在 488 nm 激发波长下获取图像。
Various fluorescent probes for the detection of intracellular reactive oxidative species (ROS) have been developed because ROS levels are closely associated with cellular states. Here, we describe a method for detection of intracellular ROS in living cells using the fluorescent probe, hydroxyphenyl fluorescein (HPF), which detects hydroxyl radicals and peroxynitrite. NIH3T3 cells andp53knockout (p53−/−) mouse embryonic fibroblasts (MEFs) were transformed by expressing oncogenic RAS using a retrovirus system. The cells were treated with HPF at 37 °C for 30 min, and subsequently, images were acquired using a confocal fluorescence microscope at an excitation wavelength of 488 nm after washing with PBS.