Electrochemiluminescent Array to Detect Oxidative Damage in ds-DNA Using [Os(bpy)2(phen-benz-COOH)]2+/Nafion/Graphene Films

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• 作者：Itti Bist ; Boya Song ; Islam M. Mosa ; Tia E. Keyes ; Aaron Martin ; Robert J. Forster ; James F. Rusling
• 刊名：ACS Sensor
• 出版年：2016
• 出版时间：March 25, 2016
• 年：2016
• 卷：1
• 期：3
• 页码：272-278
• 全文大小：642K
• ISSN：2379-3694

文摘

Reactive oxygen species (ROS) oxidize guanosines in DNA to form 8-oxo-7,8-dihydro-2-deoxyguanosine (8-oxodG), a biomarker for oxidative stress. Herein we describe a novel 64-microwell electrochemiluminescent (ECL) array enabling sensitive multiplexed detection of 8-oxodG in ds-DNA without hydrolysis. Films of Nafion and reduced graphene oxide containing ECL dye [Os(bpy)₂(phen-benz-COOH)]²⁺ (OsNG, {bpy= 2,2′-bipyridine and phen-benz-COOH = (4-(1,10-phenanthrolin-6-yl)benzoic acid)}) were assembled into microwells on a pyrolytic graphite wafer to detect 8-oxodG in oligonucleotides by electrochemiluminescence (ECL). DNA oxidation by Fenton’s reagent or by ROS formation during redox cycles involving NADPH, Cu^II, and model metabolites was monitored. UPLC-MS/MS of oxidized DNA samples were used for calibration. Detection limit for the fluidic arrays was one 8-oxodG per 670 intact nucleobases, or 0.15%. The method is sensitive enough to evaluate DNA oxidation from biologically relevant ROS-generating reactions of Cu^II, NADPH, and model metabolites.