Two-Photon Fluorescence Microscopy Imaging of Cellular Oxidative Stress Using Profluorescent Nitroxides
Ahn, Hyo-Yang , Fairfull-Smith, Kathryn E., Morrow, Benjamin J., Lussini, Vanessa, Kim, Bosung , Bondar, Mykhailo V. , Bottle, Steven E. , & Belfield, Kevin D. (2012) Two-Photon Fluorescence Microscopy Imaging of Cellular Oxidative Stress Using Profluorescent Nitroxides. Journal of the American Chemical Society, 134(10), pp. 4721-4730.
A range of varying chromophore nitroxide free radicals and their nonradical methoxyamine analogues were synthesized and their linear photophysical properties examined. The presence of the proximate free radical masks the chromophore’s usual fluorescence emission, and these species are described as profluorescent. Two nitroxides incorporating anthracene and fluorescein chromophores (compounds 7 and 19, respectively) exhibited two-photon absorption (2PA) cross sections of approximately 400 G.M. when excited at wavelengths greater than 800 nm. Both of these profluorescent nitroxides demonstrated low cytotoxicity toward Chinese hamster ovary (CHO) cells. Imaging colocalization experiments with the commercially available CellROX Deep Red oxidative stress monitor demonstrated good cellular uptake of the nitroxide probes. Sensitivity of the nitroxide probes to H2O2-induced damage was also demonstrated by both one- and two-photon fluorescence microscopy. These profluorescent nitroxide probes are potentially powerful tools for imaging oxidative stress in biological systems, and they essentially “light up” in the presence of certain species generated from oxidative stress. The high ratio of the fluorescence quantum yield between the profluorescent nitroxide species and their nonradical adducts provides the sensitivity required for measuring a range of cellular redox environments. Furthermore, their reasonable 2PA cross sections provide for the option of using two-photon fluorescence microscopy, which circumvents commonly encountered disadvantages associated with one-photon imaging such as photobleaching and poor tissue penetration.
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|Item Type:||Journal Article|
|Subjects:||Australian and New Zealand Standard Research Classification > CHEMICAL SCIENCE (030000)|
|Divisions:||Current > QUT Faculties and Divisions > Science & Engineering Faculty|
|Copyright Owner:||Copyright © 2012 American Chemical Society|
|Copyright Statement:||This is an open-access article distributed under the ACS AuthorChoice Terms & Conditions. Any use of this article, must conform to the terms of that license which are available at http://pubs.acs.org.|
|Deposited On:||12 Apr 2012 15:35|
|Last Modified:||12 Apr 2012 15:35|
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