Sensitivity of activatable reactive oxygen species probes by fluorescence spectroelectrochemistry

Abstract

We have developed a new analytical method of evaluating activatable fluorescent probes for ROS detection using integrated fluorescence spectroelectrochemistry. Tafel formalism was applied to describe the process of the probes’ oxidation under electrochemical conditions and identify a novel parameter defined as the threshold oxidation potential. This potential can serve as an approximation to the equilibrium potential and can be utilized for determining the sensitivity of a probe to oxidation. Based upon the measured values of threshold potentials, the order of sensitivity towards oxidation among several mostly used probes was determined to be following (from highest to lowest): 2,7-dichlorodihydrofluorescein > dihydroethidium > dihydrorhodamine 123 > dihydrorhodamine 6G. The presented approach opens up a new direction in synthesizing and screening novel ROS probes with a well-defined sensitivity for in vitro and in vivo applications.

Activation of the Nrf2-regulated antioxidant cell response inhibits HEMA-induced oxidative stress and supports cell viability

“…Residual superoxide anions in this group of oxidants might then be indicated by weak DHE fluorescence. However, we cannot rule out that low amounts of H 2 O 2 were formed after short exposure periods in cells treated with HEMA but remained undetected due to a relatively high threshold oxidation potential compared with DCFH 2 [33]. The level of H 2 O 2 as shown by DHR123 fluorescence was sufficiently high in HEMAexposed cell cultures after long exposure.…”

“…DCFH 2 -DA is rapidly cleaved by cellular esterases, and the resulting H 2 DCF molecule is oxidized to the fluorescent product DCF reported to be appropriate for the measurement of various ROS including peroxynitrite, hydroxyl radicals, and H 2 O 2 in combination with cellular peroxidases, although a particular reactive species was not detected [31,32]. Most relevant for the analysis of HEMA-induced ROS formation was the recent finding that DCFH 2 is characterized by a low threshold oxidation potential followed by DHE and DHR123, making DCFH 2 highly sensitive to oxidation [33].…”

Imaging of radicals following injury or acute stress in peripheral nerves with activatable fluorescent probes

“…Given the same sensitivity, probes with higher reactivity are expected to produce stronger signal. For the probe selections, quantitative measurements of sensitivity and reactivity were performed using a recently described iFOS system [26] modified for NIR measurements. In this method, electrochemical oxidation of the non-fluorescent molecules converts them into fluorescent molecules.…”

“…The sensitivity of the dye to ROS was performed spectroelectrochemically using an in-house built iFOS technique [26] modified for NIR studies with a light-emitting diode (LED) 725 nm (Thorlabs) as the source of excitation. The method combines spectroscopic measurements with an electrochemical method of oxidation to measure the threshold oxidation potential of leucodyes.…”

A formation model of superoxide radicals photogenerated in nano-TiO2 suspensions

“…In photocatalytic reactions, O 2 c À is continuously undergoing the processes of formation and deactivation simultaneously. Many methods have been developed to determine the O 2 c À , including electron spin resonance (ESR), 10 spectrophotometric assays, [11][12][13] and uorescence assays, 14,15 but the O 2 c À could only be detected at discrete times, and thus the total quantity could only be given within the irradiation duration. Given these limitations, O 2 c À dynamic monitoring is a great challenge.…”