Imaging Thiol-Based Redox Processes in Live Cells

Abstract

Biotic and abiotic stresses often lead to transient oxidation of the internal milieu of cells. This oxidation is considered to be an essential signal for further downstream events within the cellular signaling network and therefore demands careful and, if possible, quantitative analysis. Fluorescent probes and confocal or multi-photon imaging offer almost unparalleled opportunities for visualization of an ever increasing range of compounds and even the dynamics of physiological processes within living cells and tissues with minimal perturbation. However, the utility of the methods relies on the development of probes and imaging protocols that can achieve sufficient specificity and sensitivity to give unambiguous physiological measurements. In situ estimates of glutathione have been undertaken on relatively few systems so far and in vivo measurements of redox state have only recently been made possible with the advent of transgenic fluorescent reporters. Detection and quantification of reactive oxygen species (ROS) is also challenging, and a large number of different approaches have been developed to deduce information about components of this ROS-dependent signaling pathway. This review describes different imaging-based approaches for qualitative and quantitative measurement of glutathione and ROS in living plant cells. Particular attention is given to general strategies for probe design and application of quantitative confocal fluorescence imaging.