Redox homeostasis plays multiple roles in essentially all aspects of cellular function, and hence, reliable methods for measuring cellular or tissue redox status are key elements in understanding the redox related signal pathways. However, in the free radical biology field, there are many controversies on the methods to measure reactive oxygen species. In this chapter we describe our experience in measuring superoxide, hydrogen peroxide, and a general redox status using redox-sensitive green fluorescence proteins (roGFPs) in human melanoma cells.
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This study is supported by Oxnard Foundation and Waltmar Foundation to F.L.M., and Department of Medicine Allan Hubbell Education Funds (University of California Irvine) to F.L.S.
Ziech D, Franco R, Pappa A, Panayiotidis MI (2011) Reactive oxygen species (ROS)—induced genetic and epigenetic alterations in human carcinogenesis. Mutat Res 711(1–2):167–173CrossRefPubMedGoogle Scholar
Kalinina EV, Chernov NN, Saprin AN (2008) Involvement of thio-, peroxi-, and glutaredoxins in cellular redox-dependent processes. Biochemistry (Mosc) 73(13):1493–1510CrossRefGoogle Scholar
Peshavariya HM, Dusting GJ, Selemidis S (2007) Analysis of dihydroethidium fluorescence for the detection of intracellular and extracellular superoxide produced by NADPH oxidase. Free Radic Res 41(6):699–712CrossRefPubMedGoogle Scholar
Bass DA, Parce JW, Dechatelet LR, Szejda P, Seeds MC, Thomas M (1983) Flow cytometric studies of oxidative product formation by neutrophils: a graded response to membrane stimulation. J Immunol 130(4):1910–1917PubMedGoogle Scholar
Myhre O, Andersen JM, Aarnes H, Fonnum F (2003) Evaluation of the probes 2′,7′-dichlorofluorescin diacetate, luminol, and lucigenin as indicators of reactive species formation. Biochem Pharmacol 65(10):1575–1582CrossRefPubMedGoogle Scholar
Rota C, Chignell CF, Mason RP (1999) Evidence for free radical formation during the oxidation of 2′-7′-dichlorofluorescin to the fluorescent dye 2′-7′-dichlorofluorescein by horseradish peroxidase: possible implications for oxidative stress measurements. Free Radic Biol Med 27(7–8):873–881CrossRefPubMedGoogle Scholar
Cannon MB, Remington SJ (2008) Redox-sensitive green fluorescent protein: probes for dynamic intracellular redox responses. A review. Methods Mol Biol 476:51–65PubMedGoogle Scholar
Hanson GT, Aggeler R, Oglesbee D, Cannon M, Capaldi RA, Tsien RY, Remington SJ (2004) Investigating mitochondrial redox potential with redox-sensitive green fluorescent protein indicators. J Biol Chem 279(13):13044–13053CrossRefPubMedGoogle Scholar
Dooley CT, Dore TM, Hanson GT, Jackson WC, Remington SJ, Tsien RY (2004) Imaging dynamic redox changes in mammalian cells with green fluorescent protein indicators. J Biol Chem 279(21):22284–22293CrossRefPubMedGoogle Scholar
1.Department of Epidemiology, Chao Family Comprehensive Cancer CenterUniversity of California Irvine School of MedicineIrvineUSA
2.Beckman Laser Institute and Medical ClinicUniversity of California IrvineIrvineUSA
3.The State Key Laboratory of Medical Genetics and School of Life SciencesCentral South UniversityChangsha, Hunan ProvinceChina
4.Institute of Mitochondrial Biology and Medicine, The Key Laboratory of Biomedical Information Engineering of Ministry of EducationXi’an Jiaotong University School of Life Science and TechnologyXi’an, Shaangxi ProvinceChina
5.Department of Medicine, Chao Family Comprehensive Cancer CenterUniversity of California Irvine School of MedicineIrvineUSA