Abstract
Reactive oxygen species (ROS) exhibit different spatial and temporal distributions as well as concentrations in- and outside the cell, thereby functioning as signaling or pathogen-destroying molecules. Especially the ROS H2O2 is important for the patho/physiological status of an organism. Electrochemistry (EM) and electron spin resonance (ESR)-based techniques allow quantification of H2O2 in artificial and living systems, coping a concentration range from low nM up to mM. Working electrodes for EM are optimized by diverse modifications and, additionally, redox mediators are used. Ultramicroelectrodes allow scanning of single cells to spatially resolve and quantify extracellular H2O2 in real-time. With ESR spectroscopy, •O2¯, but not H2O2, can be directly determined by spin probes in- and outside of cells in suspensions. Monitoring H2O2 requires formation of intermediate radicals, detectable with spin probes. Low μM [H2O2] can thus be assessed specifically. Using suitable spin traps, in-vivo ESR and immuno-spin trapping can visualize different radicals at their respective production sites in small animals, organs and tissues. Here, the redox reaction cascades may interfere with cell metabolism. Optimization of all methods established for H2O2 determination would be favorable to finally combine them for mutual validation. Thus, a deeper insight into cellular ROS metabolism can be obtained.
Highlights
Hydrogen Peroxide (H2O2) is widely recognized as one of the most important small molecules responsible for physiological functioning of cells in prokaryotes as well as eukaryotes.[1]
H2O2 is formed by a sequential 2etransfer to O2, whereby the intermediate radical O2 ̄ is produced. O2 ̄ is subsequently protonated resulting in the nonradical H2O2
Oxygen reduction under physiological conditions is controlled by special enzymes which contribute to a balanced reactive oxygen species (ROS) level inside and outside the cell.[13]
Summary
Hydrogen Peroxide (H2O2) is widely recognized as one of the most important small molecules responsible for physiological functioning of cells in prokaryotes as well as eukaryotes.[1]. Diverse approaches to measure ROS in organisms, tissues and cells have been developed in the last decades Some of these methods are based on enzymatic, fluorescent and luminescent techniques.[65,66,67,68,69,70,71] More recent approaches use genetically-encoded protein sensors or transgenic organisms to determine H2O2.72–79 While these approaches all have advantages and disadvantages,[65,71,80,81,82,83,84] one of their major problems is the quantification of absolute [H2O2] or even cellular [H2O2] kinetics.
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