Capillary Electrophoresis of Oxidative Dna Damage

Xu, Guowang; Shi, Xianzhe; Mei, Surong; Yao, Qinghong; Weng, Qianfeng; Wu, Caiying

doi:10.1007/978-1-59745-376-9_16

Cited by 4 publications

(3 citation statements)

References 16 publications

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“…Furthermore, it is known that interactions between DNA and ROS produce DNA strand break and base modification, which are frequently assessed by measurement of 8-OHdG levels . Interestingly, a recent study demonstrates that urinary 8-OHdG is an excellent marker for oxidative DNA damage (Xu et al 2008).…”

Section: Discussionmentioning

confidence: 99%

Glial Damage After Transient Focal Cerebral Ischemia in Rats

et al. 2008

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We investigated the immunohistochemical changes of 8-hydroxy-2'-deoxyguanosine (8-OHdG) immunoreactivity as a marker of DNA damage and single-strand DNA (ssDNA) immunoreactivity as a marker of apoptosis in the striatum from 1 up to 15 days after 90 min of focal cerebral ischemia caused by middle cerebral artery occlusion in rats. In the present study, marked loss of MAP2 immunostaining was observed in the ipsilateral striatum 3 days after focal cerebral ischemia. A significant increase in the number of ssDNA-immunoreactive apoptotic neurons was observed in the ipsilateral striatum 1 and 3 days after focal cerebral ischemia. In contrast, a significant increase in densities of 8-OHdG-immunopositive cells was observed in the ipsilateral striatum from 3 up to 15 days after focal cerebral ischemia. Our double-labeled immunochemical study showed that 8-OHdG immunoreactivity was observed in both isolectin B(4)-positive microglia and glial fibrillary acidic protein-immunopositive astrocytes in the ipsilateral striatum 7 days after focal cerebral ischemia. These results suggest that focal cerebral ischemia can cause a marked increase in the number of microglia and astrocytes with oxidative DNA damage in the ipsilateral striatum. Furthermore, our results show that most microglia and astrocytes in the ipsilateral striatum after focal cerebral ischemia may not die by apoptosis. Thus, our findings provide novel evidence that focal cerebral ischemia can cause oxidative DNA damage in most microglia and astrocytes.

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Section: Discussionmentioning

confidence: 99%

Glial Damage After Transient Focal Cerebral Ischemia in Rats

et al. 2008

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“…capillaries or smaller, it has been found that the voltage drop across the capillary is great enough that interference with the detection circuit is minimal [17]; however, a shift in the half-wave potential is possible [18]. This configuration has been used for high-throughput separations [19], analysis of insecticides [20], and quantitation of DNA damage [21].…”

Section: Introductionmentioning

confidence: 99%

The Use of a 3D‐printed Microfluidic Device and Pressure Mobilization for Integrating Capillary Electrophoresis with Electrochemical Detection

Kimlinger

Martin

2018

Electroanalysis

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Capillary electrophoresis coupled with electrochemical detection can be a powerful analysis tool; however, previous methods developed to integrate these two techniques can often times be fragile and have alignment issues such that there are no commercially available approaches. In this paper, we present the use of a 3D-printed Wall-Jet Electrode device for integrating capillary electrophoresis with electrochemical detection. A pressure mobilization step was also utilized to further reduce noise by allowing the electrophoresis separation step to continue only until the first analyte was close to elution. Then, the separation voltage was terminated and pressure-based flow was used for elution of the analyte bands onto the electrode surface with a wall-jet configuration. It is shown that the pressure-based elution is beneficial for the reduction of baseline noise and elimination of field effects. A mixture of catecholamines were separated to demonstrate effectiveness of the system. In addition, the system was coupled with a Beckman Coulter commercial capillary electrophoresis instrument in a straightforward manner. The system was also shown to be effective in separations done with a high ionic strength physiological buffer. This 3D printing approach can be used by researchers to utilize electrochemical detection on commercial capillary electrophoresis systems by downloading the provided STL and/or CAD files.

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“…Capillary electrophoresis with UV or electrochemical detection has been used to analyze the oxidative damage of DNA. However, these assays exhibited low sensitivity [20]. In addition, capillary electrophoresis coupled to laser-induced fluorescence (CE-LIF) was developed to sensitively measure endogenous damage in mitochondrial and genomic DNA [21].…”

Section: Introductionmentioning

confidence: 99%

Selective enzymatic cleavage and labeling for sensitive capillary electrophoresis laser-induced fluorescence analysis of oxidized DNA bases

Wang

2015

Journal of Chromatography A

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a b s t r a c tOxidatively generated DNA damage is considered to be a significant contributing factor to cancer, aging, and age-related human diseases. It is important to detect oxidatively generated DNA damage to understand and clinically diagnosis diseases caused by oxidative damage. In this study, using selective enzymatic cleavage and quantum dot (QD) labeling, we developed a novel capillary electrophoresislaser induced fluorescence method for the sensitive detection of oxidized DNA bases. First, oxidized DNA bases are recognized and removed by one DNA base excision repair glycosylase, leaving apurinic and apyrimidinic sites (AP sites) at the oxidized positions. The AP sites are further excised by the AP nicking activity of the chosen glycosylase, generating a nucleotide gap with 5 -and 3 -phosphate groups. After dephosphorylation with one alkaline phosphatase, a biotinylated ddNTP is introduced into the nucleotide space within the DNA strand by DNA polymerase I. The biotin-tagged DNA is further labeled with a QD-streptavidin conjugate via non-covalent interactions. The DNA-bound QD is well-separated from excess DNA-unbound QD by highly efficient capillary electrophoresis and is sensitively detected by online coupled laser-induced fluorescence analysis. Using this method, we can assess the trace levels of oxidized DNA bases induced by the Fenton reaction and UV irradiation. Interestingly, the use of the formamidopyrimidine glycosylase (FPG) protein and endonuclease VIII enables the detection of oxidized purine and pyrimidine bases, respectively. Using the synthesized standard DNA, the approach has low limits of detection of 1.1 × 10 −19 mol in mass and 2.9 pM in concentration.

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Capillary Electrophoresis of Oxidative Dna Damage

Cited by 4 publications

References 16 publications

Glial Damage After Transient Focal Cerebral Ischemia in Rats

Glial Damage After Transient Focal Cerebral Ischemia in Rats

The Use of a 3D‐printed Microfluidic Device and Pressure Mobilization for Integrating Capillary Electrophoresis with Electrochemical Detection

Selective enzymatic cleavage and labeling for sensitive capillary electrophoresis laser-induced fluorescence analysis of oxidized DNA bases

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