Ultrasensitive nuclease activity and inhibition assay using microchip electrophoresis with laser induced fluorescence detection

Qin, Yingfeng; Zhao, Jingjin; Huang, Yong; Li, Shuting; Zhao, Shulin

doi:10.1039/c5ay02831k

Cited by 9 publications

(4 citation statements)

References 43 publications

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Section: Methods and Techniques To Quantify Nuclease-mediated Degrada...mentioning

confidence: 99%

“…After separation of the different fragments with CE, the fluorescence intensity of 5-FAM-nucleoside monophosphates was used to quantify the nuclease activity and the effect of nuclease inhibitors. 374 CE is highly efficient, versatile, reproducible, ease-to-use and requires low sample and buffer volume to quantify and analyze the length distribution of nucleic acids. The combination of CE with advanced detection techniques like mass spectroscopy, is becoming an indispensable analytical tool to separate and quantify nucleic acids.…”

Section: Capillary Electrophoresis (Ce)mentioning

confidence: 99%

See 1 more Smart Citation

Nucleic acid degradation as barrier to gene delivery: a guide to understand and overcome nuclease activity

Zhang,

Vandesompele,

Braeckmans

et al. 2024

Chem. Soc. Rev.

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Section: Methods and Techniques To Quantify Nuclease-mediated Degrada...mentioning

confidence: 99%

Section: Capillary Electrophoresis (Ce)mentioning

confidence: 99%

Nucleic acid degradation as barrier to gene delivery: a guide to understand and overcome nuclease activity

Zhang,

Vandesompele,

Braeckmans

et al. 2024

Chem. Soc. Rev.

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“…Comparing with normal capillary zone electrophoresis mode, it achieved the detection sensitivity enhancement and improved from 61 to 265 folds. Fluorescein‐labeled substrate enzyme assay is an effective approach for improving enzyme activity detection and inhibitor screening . A microchip electrophoresis method with LIF for ultrasensitive detection of nuclease activity and inhibitors was reported .…”

Section: Off‐line Screening Of Enzyme Inhibitorsmentioning

confidence: 99%

“…Fluorescein‐labeled substrate enzyme assay is an effective approach for improving enzyme activity detection and inhibitor screening . A microchip electrophoresis method with LIF for ultrasensitive detection of nuclease activity and inhibitors was reported . In the assay, the fluorescein‐labeled ssDNA (FAM‐ssDNA) would be digested to 5‐FAM‐nucleoside monophosphates and short oligonucleotide fragments by nuclease.…”

Section: Off‐line Screening Of Enzyme Inhibitorsmentioning

confidence: 99%

Advances in screening enzyme inhibitors by capillary electrophoresis

Wang

Yang

2019

Electrophoresis

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Capillary electrophoresis (CE) has attracted lots of attention due to its simplicity, low sample consumption, low solvent volume, high resolution, and high speed. Based on these advantages, it has been widely used in enzyme inhibitor screening. There are two main operation modes on enzyme inhibitor screening: off‐line (precapillary enzyme assays) in which process CE was used as an analytical tool; online (in‐capillary enzyme assays) which combined the sample injection, mix, reaction, separation, and detection within a single run. Additionally, diverse of new materials were introduced to immobilize enzyme, which has been coupled with CE for the study of enzyme activity and its inhibitor screening. This review gives an overview of the developments and applications for the CE‐based enzyme inhibitor screening.

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A review on microscale polymerase chain reaction based methods in molecular diagnosis, and future prospects for the fabrication of fully integrated portable biomedical devices

Lee

2018

Microchim Acta

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Since the advent of microfabrication technology and soft lithography, the lab-on-a-chip concept has emerged as a state-of-the-art miniaturized tool for conducting the multiple functions associated with micro total analyses of nucleic acids, in series, in a seamless manner with a miniscule volume of sample. The enhanced surface-to-volume ratio inside a microchannel enables fast reactions owing to increased heat dissipation, allowing rapid amplification. For this reason, PCR has been one of the first applications to be miniaturized in a portable format. However, the nature of the basic working principle for microscale PCR, such as the complicated temperature controls and use of a thermal cycler, has hindered its total integration with other components into a micro total analyses systems (μTAS). This review (with 179 references) surveys the diverse forms of PCR microdevices constructed on the basis of different working principles and evaluates their performances. The first two main sections cover the state-of-the-art in chamber-type PCR microdevices and in continuous-flow PCR microdevices. Methods are then discussed that lead to microdevices with upstream sample purification and downstream detection schemes, with a particular focus on rapid on-site detection of foodborne pathogens. Next, the potential for miniaturizing and automating heaters and pumps is examined. The review concludes with sections on aspects of complete functional integration in conjunction with nanomaterial based sensing, a discussion on future prospects, and with conclusions. Graphical abstract In recent years, thermocycler-based PCR systems have been miniaturized to palm-sized, disposable polymer platforms. In addition, operational accessories such as heaters and mechanical pumps have been simplified to realize semi-automatted stand-alone portable biomedical diagnostic microdevices that are directly applicable in the field. This review summarizes the progress made and the current state of this field.

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Ultrasensitive nuclease activity and inhibition assay using microchip electrophoresis with laser induced fluorescence detection

Abstract: A novel microchip electrophoresis method with laser induced fluorescence detection was developed for ultrasensitive detection of nuclease activity and inhibitors.

Cited by 9 publications

References 43 publications

Nucleic acid degradation as barrier to gene delivery: a guide to understand and overcome nuclease activity

Nucleic acid degradation as barrier to gene delivery: a guide to understand and overcome nuclease activity

Advances in screening enzyme inhibitors by capillary electrophoresis

A review on microscale polymerase chain reaction based methods in molecular diagnosis, and future prospects for the fabrication of fully integrated portable biomedical devices

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