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Description:
Provides all necessary reagents, buffers and a detailed protocol for the detection of Reactive Oxygen Species (ROS, peroxidase activity) in live cells.
Application:
Fluorogenic peroxidase substrates are converted to highly fluorescent products in the presence of the enzyme and hydrogen peroxide but can be relatively unstable for use in enzyme-linked immunosorbent assays (ELISAs). By forming the diacetate derivates, intracellular applications are more efficient, whereupon the acetates are cleaved by endogenous esterases, releasing the intact substrate. This assay utilizes the cell permeable reagent, 2,7-dichlorofluorescin diacetate M0807, a reliable fluorogenic marker for ROS detection. Upon enzyme activity, the highly fluorescent dye, 2,7-dichlorofluorescein is produced, with EX: 495nm and EM: 529 nm.
References:
View References ▼
- Lyons TA, Amouretti XF, Held PG, Naleway JJ (2010) "Development of a Live-Cell Based Reactive Oxygen Species (ROS) Assay for use in High-Content Screening of Drug Candidates Using the BioTek Synergy Mx Microplate Reader." presented at the Society for Biomolecular Sciences 16th Annual Conference in Phoenix, AZ April 11-15, 2010. link
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Kitabchi AE, Stentz FB, Umpierrez GE. (2004) "Diabetic ketoacidosis induces in vivo activation of human T-lymphocytes." Biochem Biophys Res Commun 315: 404-7.
- Kumar SS, Shankar B, Sainis KB. (2004)"Effect of chlorophyllin against oxidative stress in splenic lymphocytes in vitro and in vivo." Biochim Biophys Acta 1672: 100-11.
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- Delogu G, Antonucci A, Moretti S, Marandola M, Tellan G, Signore M, Famularo G. (2004) "Oxidative stress and mitochondrial glutathione in human lymphocytes exposed to clinically relevant anesthetic drug concentrations." J Clin Anesth 16: 189-94.
- Kalivendi S.V., Kotamraju S., Cunningham S., Shang T., Hillard C.J., Kalyanaraman B. (2003) “1-Methyl-4-phenylpyridinium (MPP+)-induced apoptosis and mitochondrial oxidant generation: role of transferrin-receptor-dependent iron and hydrogen peroxide.” Biochem J 371:151-64.
- Chen CY, Wang YF, Lin YH, Yen SF. (2003) "Nickel-induced oxidative stress and effect of antioxidants in human lymphocytes." Arch Toxicol 77: 123-30.
- Diaz G, Liu S, Isola R, Diana A, Falchi AM. (2003) "Mitochondrial localization of reactive oxygen species by dihydrofluorescein probes." Histochem. Cell Biol. 120: 319-25 .
- Tampo Y, Kotamraju S, Chitambar CR, Kalivendi SV, Keszler A, Joseph J, Kalyanaraman B. (2003) "Oxidative stress-induced iron signaling is responsible for peroxide-dependent oxidation of dichlorodihydrofluorescein in endothelial cells: role of transferrin receptor-dependent iron uptake in apoptosis." Circ. Res. 92: 56-63.
- Blattner JR, He L, Lemasters JJ. (2001) "Screening assays for the mitochondrial permeability transition using a fluorescence multiwell plate reader." Anal. Biochem. 295: 220-226.
- Kehrer JP, Paraidathathu T (1992) "The use of fluorescent probes to assess oxidative processes in isolated-perfused rat heart tissue." Free Radic. Res. Commun. 16: 217-222.
- Salata RA, Sullivan JA, Mandell GL. (1983) "Visualization of hydrogen peroxide in living polymorphonuclear neutrophils utilizing leucodiacetyl 2,7-dichlorofluorescin: photomicrographic and microphotometric studies." Trans. Assoc. Am. Physicians 96: 375-383.
- Keston AS, Brandt R (1965)"The fluorometric analysis of ultramicro quantities of hydrogen peroxide." Anal. Biochem. 11: 1-5.
- Lee TK, O’Brien KF, Wang W, Johnke RM,
Sheng C, Benhabib SM, Wang T, Allison RR. (2010) “Radioprotective effect of American ginseng
on human lymphocytes at 90 Minutes postirradiation:
A study of 40 cases.” J Altern Complement Med 16:561-567.
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Unit Price (USD):
1 kit $211.02
Bulk Price/unit
(5 or more): $168.82
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