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Imaging the Redox States of Live Cells with the Time-Resolved Fluorescence of Genetically Encoded Biosensors

Li, Lei, Zhang, Changcheng, Wang, Peng, Wang, Aoxue, Zhou, Jiasheng, Chen, Guoqing, Xu, Jianhua, Yang, Yi, Zhao, Yuzheng, Zhang, Sanjun, Tian, Yang
Analytical chemistry 2019 v.91 no.6 pp. 3869-3876
antineoplastic agents, biosensors, fluorescence, fluorescent proteins, image analysis, macrophages, mitochondria, oxidative stress, thiols, vertebrates, wavelengths
Redox environments in cells influence many important physiological and pathological processes. In this study, the time-resolved fluorescence of a recently reported thiol redox-sensitive sensor based on vertebrate fluorescent protein UnaG, roUnaG, was studied, along with the application of the time-resolved fluorescence of roUnaG to image the redox states of the mitochondria, cytoplasm, and nucleus in live cells. Time-resolved fluorescence images of roUnaG clearly demonstrated that potent anticancer compound KP372-1 induced extreme oxidative stress. A more stressful redox state observed in activated macrophages further demonstrated the validity of roUnaG with time-resolved fluorescence. For comparison, time-resolved fluorescence images of four other frequently used redox biosensors (roGFP1, HyPer, HyPerRed, and rxRFP) were also captured. The time-resolved fluorescence allows an intrinsically ratiometric measurement for biosensors with one excitation wavelength and provides new opportunities for bioimaging.