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

Bist, Itti; Song, Boya; Mosa, Islam M.; Keyes, Tia E.; Martin, Aaron; Forster, Robert J.; Rusling, James F.

doi:10.1021/acssensors.5b00189

Cited by 32 publications

(43 citation statements)

References 53 publications

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“…56,57 In the presence of Cu 2+ and an enzymatic NADPH regenerating system, redox cycling of catechol, semiquinone radical, and benzoquinone produces ROS, which oxidize dG to 8-oxodG in DNA (Scheme 1). 15,16,58 …”

Section: Resultsmentioning

confidence: 99%

“…Conditions were optimized to limit G oxidations to 8-oxodG without strand breaks and to yield LC-MS/MS results of sufficient intensity for oxidation site location and quantitation. Briefly, 100 μ g of oligonucleotide was incubated in 200 μ L of Tris buffer with 1 mM catechol, 15 50 μ M CuCl 2 , and NADPH regeneration system (10 mM G6P, one unit G6PDH enzyme, 0.8 mM NADP + , 1 mM MgCl 2 ) at 37 °C for 12 h. Alternatively, 100 μ g of oligonucleotide was incubated with Fenton’s reagent containing 0.1 mM FeSO 4 and 40 mM H 2 O 2 16 at 37 °C for 4 h with constant stirring. Amicon Ultra-0.5 3K centrifugal filters from MilliporeSigma were used to remove excess catechol, NADPH system, and FeSO 4 products.…”

Section: Methodsmentioning

confidence: 99%

“…ROS, including H 2 O 2 , O 2 •− , and • OH, are formed in these redox cycles (Scheme 1) and can oxidize DNA in vitro. 15,16 Also, polycyclic aromatic hydrocarbon (PAH) o -quinones are implicated as causes of mutations in tumor suppressor genes and are known risk factors for lung cancer. 17 …”

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confidence: 99%

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Direct LC-MS/MS Detection of Guanine Oxidations in Exon 7 of the p53 Tumor Suppressor Gene

Jiang

Malla

et al. 2017

Anal. Chem.

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Oxidation of DNA by reactive oxygen species (ROS) yields 8-oxo-7,8-dihydroguanosine (8-oxodG) as primary oxidation product, which can lead to downstream G to T transversion mutations. DNA mutations are nonrandom, and mutations at specific codons are associated with specific cancers, as widely documented for the p53 tumor suppressor gene. Here, we present the first direct LC-MS/MS study (without isotopic labeling or hydrolysis) of primary oxidation sites of p53 exon 7. We oxidized a 32 base pair (bp) double-stranded (ds) oligonucleotide representing exon 7 of the p53 gene. Oxidized oligonucleotides were cut by a restriction endonuclease to provide small strands and enable positions and amounts of 8-oxodG to be determined directly by LC-MS/MS. Oxidation sites on the oligonucleotide generated by two oxidants, catechol/Cu2+/NADPH and Fenton’s reagent, were located and compared. Guanines in codons 243, 244, 245, and 248 were most frequently oxidized by catechol/Cu2+/NADPH with relative oxidation of 5.6, 7.2, 2.6, and 10.7%, respectively. Fenton’s reagent oxidations were more specific for guanines in codons 243 (20.3%) and 248 (10.4%). Modeling of docking of oxidizing species on the ds-oligonucleotide were consistent with the experimental codon oxidation sites. Significantly, codons 244 and 248 are mutational “hotspots” in nonsmall cell and small cell lung cancers, supporting a possible role of oxidation in p53 mutations leading to lung cancer.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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Direct LC-MS/MS Detection of Guanine Oxidations in Exon 7 of the p53 Tumor Suppressor Gene

Jiang

Malla

et al. 2017

Anal. Chem.

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“…Recently, the oxidation of guanosines in DNA from the reactive oxygen species (ROS) was evaluated by using a new [Os(bpy) 2 (phen-benz-COOH)] 2? /nafion/graphene film [98].…”

Section: Genotoxicity Screeningmentioning

confidence: 99%

Imaging Analysis Based on Electrogenerated Chemiluminescence

et al. 2017

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Electrochemiluminescence (ECL), also called electrogenerated chemiluminescence, is luminescence that is produced by chemical reactions triggered by electrochemical method. ECL imaging is a novel imaging technique, which can simultaneously provide two kinds of signals of both electrochemistry and optical image. Over the past two decades, ECL imaging has been not only a powerful tool for investigating the fundamental scientific questions such as ECL mechanisms and reaction kinetics, but also a versatile analytical technique for detection of a wide range of analytes including small molecules, DNA, proteins, and cells. In the first part of this review, we briefly describe the reaction mechanisms of ECL generation. Then the review focuses on the research progress on the ECL imaging approach. It is basically introduced from the following five aspects: (i) visualization of electroactive sites on surfaces, (ii) imaging analysis at the single-bead and single-cell levels, (iii) array bioanalysis, (iv) multi-color ECL imaging, and (v) paper chip based on ECL imaging. Finally, some perspectives and future directions in this active research area are presented.

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“…In a different approach, osmium tetroxide-2,2 -bipyridine was employed as an electrochemical probe of AP sites, which reacts covalently with the unpaired thymine bases in damaged DNA [23]. Furthermore, although a few chemical toxicity sensors based on the detection of oxidized or methylated DNA bases have been developed [24][25][26][27][28][29][30][31][32][33][34][35][36], so far there has been no report on the study of toxicity sensors based on AP site detection.…”

Section: Introductionmentioning

confidence: 99%

A chemical toxicity sensor based on the electrochemiluminescence quantification of apurinic/apyrimidinic sites in double-stranded DNA monolayer

Feng

Liang

Guo

et al. 2017

Sensors and Actuators B: Chemical

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a b s t r a c tA new chemical toxicity sensor was developed based on the electrochemiluminescence (ECL) quantification of apurinic/apyrimidinic sites (AP sites) in a DNA monolayer with a covalent aldehyde reactive probe (ARP). In the sensor, a uracil-containing DNA duplex was first immobilized on a gold electrode by self-assembly. The duplex was then reacted with uracil-DNA glycosylase (UDG) to convert uracils into AP sites. ARP was employed to tag the AP site with a biotin. After reacting with a ruthenium complex labeled streptavidin, ECL was measured for quantitative analysis. The DNA monolayer was characterized by cyclic voltammetry, electrochemical impedance spectroscopy and chronocoulometry, and its density was measured to be 2.89 × 10 −12 mol/cm 2 . Characterization of the reaction product between ARP and DNA AP sites in solution by nondenaturing polyacrylamide gel electrophoresis and mass spectrometry confirmed successful biotinylation. ECL intensity of the labeled DNA monolayer on the electrode was found to correlate with the number of AP sites, and the detection limit was estimated to be about 1 lesion in 512 DNA bases, which meant that 8.5 fmol AP bases on the electrode were detected. ECL response of the DNA monolayers containing either 8-oxodGuo or methylated bases was very low, indicating that ARP-based AP sites detection method was highly selective. The sensor successfully detected the AP sites in normal DNA induced by methylmethane sulfonate, a carcinogenic chemical. The novel combination of covalent probe and ECL measurement in a sensor configuration therefore provides unique advantages in selectivity and sensitivity, and can be potentially employed in the screening of chemicals for their genetic toxicity.

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

Cited by 32 publications

References 53 publications

Direct LC-MS/MS Detection of Guanine Oxidations in Exon 7 of the p53 Tumor Suppressor Gene

Direct LC-MS/MS Detection of Guanine Oxidations in Exon 7 of the p53 Tumor Suppressor Gene

Imaging Analysis Based on Electrogenerated Chemiluminescence

A chemical toxicity sensor based on the electrochemiluminescence quantification of apurinic/apyrimidinic sites in double-stranded DNA monolayer

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

Cited by 32 publications

References 53 publications

Direct LC-MS/MS Detection of Guanine Oxidations in Exon 7 of the p53 Tumor Suppressor Gene

Direct LC-MS/MS Detection of Guanine Oxidations in Exon 7 of the p53 Tumor Suppressor Gene

Imaging Analysis Based on Electrogenerated Chemiluminescence

A chemical toxicity sensor based on the electrochemiluminescence quantification of apurinic/apyrimidinic sites in double-stranded DNA monolayer

Contact Info

Product

Resources

About

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