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10 May 1996 Imaging of hydrogen peroxide generation in cultured cells using dichlorofluorescein derivatives
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There is growing evidence that generation of reactive oxygen molecules (e.g., hydrogen peroxide, superoxide, hydroxyl and nitric oxide radicals) plays an important role in cell death. In this report we evaluated the effectiveness of the membrane permeable probe carboxy-dichlorodihydrofluorescein diacetate acetoxymethyl ester (C-DCDHF-DA-AM) for imaging the production of H2O2 in cultured cells. We examined the properties of three derivatives of the ester in saline droplets and compared the results with responses recorded from cells loaded with the ester. Results indicated that fluorescence was generated in cells and droplets by a photo-oxidative process involving H2O2. Videomicroscopy demonstrated that the cellular responses originated in small vesicles (presumably peroxisomes), with large responses filling the cytosol and enveloping the nucleus. We interpreted these responses as due to light-induced activation of flavin-containing oxidases, which generate H2O2 in peroxisomes, followed by diffusion of H2O2 throughout the cell. Escape of H2O2 from peroxisomes into Fe2+-containing compartments could have dire consequences on cell viability due to the production of hydroxyl free radicals. Such a mechanism could underly the phototoxic effects of visible light on cultured cells.
© (1996) COPYRIGHT Society of Photo-Optical Instrumentation Engineers (SPIE). Downloading of the abstract is permitted for personal use only.
Philip E. Hockberger, Mohammed S. Ahmed, Timothy A. Skimina, Christopher Lee, Wu-Yen Hung, and Tippu Siddique "Imaging of hydrogen peroxide generation in cultured cells using dichlorofluorescein derivatives", Proc. SPIE 2678, Optical Diagnostics of Living Cells and Biofluids, (10 May 1996);

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