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Updated: Apr 4, 2026
Quantification of Reactive Oxygen Species Using 2′,7′-Dichlorofluorescein Diacetate Probe and Flow-Cytometry in Müller Glial Cells
Published on: May 13, 2022
In Vivo Imaging of Retinal Oxidative Stress Using a Reactive Oxygen Species-Activated Fluorescent Probe
Megan C Prunty1, Moe H Aung1, Adam M Hanif2
1Department of Ophthalmology, Emory University, Atlanta, Georgia, United States.
Purpose:
In vivo methods for detecting oxidative stress in the eye would improve screening and monitoring of the leading causes of blindness: diabetic retinopathy, glaucoma, and age-related macular degeneration.
Methods:
To develop an in vivo biomarker for oxidative stress in the eye, we tested the efficacy of a reactive oxygen species (ROS)-activated, near-infrared hydrocyanine-800CW (H-800CW) fluorescent probe in light-induced retinal degeneration (LIRD) mouse models. After intravitreal delivery in LIRD rats, fluorescent microscopy was used to confirm that the oxidized H-800CW appeared in the same retinal layers as an established ROS marker (dichlorofluorescein).
Results:
Dose-response curves of increasing concentrations of intravenously injected H-800CW demonstrated linear increases in both intensity and total area of fundus hyperfluorescence in LIRD mice, as detected by scanning laser ophthalmoscopy. Fundus hyperfluorescence also correlated with the duration of light damage and functional deficits in vision after LIRD. In LIRD rats with intravitreal injections of H-800CW, fluorescent labeling was localized to photoreceptor inner segments, similar to dichlorofluorescein.
Conclusions:
Hydrocyanine-800CW detects retinal ROS in vivo and shows potential as a novel biomarker for ROS levels in ophthalmic diseases.
