On the use of fluorescence lifetime imaging and dihydroethidium to detect superoxide in intact animals and ex vivo tissues: A reassessment |
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Affiliation: | 1. Department of Biophysics and Free Radical Research Center, Medical College of Wisconsin, Milwaukee, WI 53226, USA;2. Institute of Applied Radiation Chemistry, Lodz University of Technology, 90-924 Lodz, Poland;1. Department of Chemistry, College of Science, University of Tikrit, Tikrit, Iraq;2. Department of Chemistry, College of Education for Pure Science, University of Tikrit, Tikrit, Iraq;3. Department of Nursing, Al-Hadi University College, Baghdad, Iraq;4. Center of Excellence on Solar Cells & Renewable Energy, Department of Physics, School of Basic Sciences and Research, Sharda University, Great er Noida, Uttar Pradesh 201308, India;1. Department of Pathology, Columbia University, NY, NY 10032, USA;2. Dipartimento di Medicina Sperimentale e Clinica, Università degli Studi di Firenze, Firenze 50134, Italy;1. Department of Endocrinology, Institute of Zoology, Jagiellonian University, Krakow, Poland;2. Department of Cell Biology, Poznan University of Medical Sciences, Poznan, Poland;1. Key Laboratory for Advanced Materials and Institute of Fine Chemicals, East China University of Science & Technology, Shanghai 200237, PR China;2. School of Chemical Biology and Pharmaceutical Science, Capital Medical University, Beijing 100069, PR China;1. Whitehead Institute for Biomedical Research, 9 Cambridge Center, Cambridge, MA 02142, USA;2. Department of Biology, Massachusetts Institute of Technology (MIT), Cambridge, MA 02139, USA;3. Broad Institute, Seven Cambridge Center, Cambridge, MA 02142, USA;4. David H. Koch Institute for Integrative Cancer Research at MIT, 77 Massachusetts Avenue, Cambridge, MA 02139, USA;5. Howard Hughes Medical Institute, MIT, Cambridge, MA 02139, USA;6. Department of Cell Biology, New York University School of Medicine, New York, NY 10016, USA;7. Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA 94305, USA;8. Department of Biochemistry, Netherlands Cancer Institute, Plesmanlaan 121 1066 CX, Amsterdam, the Netherlands |
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Abstract: | Recently, D.J. Hall et al. reported that ethidium (E+) is formed as a major product of hydroethidine (HE) or dihydroethidium reaction with superoxide (O2−) in intact animals with low tissue oxygen levels (J. Cereb. Blood Flow Metab. 32:23–32, 2012). The authors concluded that measurement of E+ is an indicator of O2− formation in intact brains of animals. This finding is in stark contrast to previous reports using in vitro systems showing that 2-hydroxyethidium, not ethidium, is formed from the reaction between O2− and HE. Published in vivo results support the in vitro findings. In this study, we performed additional experiments in which HE oxidation products were monitored under different fluxes of O2−. Results from these experiments further reaffirm our earlier findings (H. Zhao et al., Free Radic. Biol. Med. 34:1359, 2003). We conclude that whether in vitro or in vivo, E+ measured by HPLC or by fluorescence lifetime imaging is not a diagnostic marker product for O2− reaction with HE. |
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Keywords: | 2-hydroxyethidium DE" },{" #name" :" keyword" ," $" :{" id" :" key0020" }," $$" :[{" #name" :" text" ," _" :" deuteroethidine DHE" },{" #name" :" keyword" ," $" :{" id" :" key0030" }," $$" :[{" #name" :" text" ," _" :" dihydroethidium dtpa" },{" #name" :" keyword" ," $" :{" id" :" key0040" }," $$" :[{" #name" :" text" ," _" :" diethylenetriaminepentaacetate ethidium HE" },{" #name" :" keyword" ," $" :{" id" :" key0060" }," $$" :[{" #name" :" text" ," _" :" hydroethidine HX" },{" #name" :" keyword" ," $" :{" id" :" key0070" }," $$" :[{" #name" :" text" ," _" :" hypoxanthine X" },{" #name" :" keyword" ," $" :{" id" :" key0080" }," $$" :[{" #name" :" text" ," _" :" xanthine XO" },{" #name" :" keyword" ," $" :{" id" :" key0090" }," $$" :[{" #name" :" text" ," _" :" xanthine oxidase HRP" },{" #name" :" keyword" ," $" :{" id" :" key0100" }," $$" :[{" #name" :" text" ," _" :" horseradish peroxidase TFA" },{" #name" :" keyword" ," $" :{" id" :" key0110" }," $$" :[{" #name" :" text" ," _" :" trifluoroacetic acid |
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