2,4‐Dinitrobenzenesulfonyl Fluoresceins as Fluorescent Alternatives to Ellman's Reagent in Thiol‐Quantification Enzyme Assays

Maeda, Hatsuo; Matsuno, Hiromi; Ushida, Mai; Katayama, Kohei; Saeki, Kanako; Itoh, Norio

doi:10.1002/ange.200500114

Cited by 88 publications

(47 citation statements)

References 23 publications

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“…The majority of the reported methods are based on redox chemistry or labeling with chromophores or fluorophores and a combination of separation techniques. [7] Recently, Strongin et al as well as several others have made pioneering advances in developing highly selective probes for Cys and Hcy, as well as a thiolquantification enzyme assay based on the covalent interaction between the probe molecule and the analyte, [8,9] which could be used for the direct assay of the amino acid content and the enzyme activity in body fluids. In addition, a sensing ensemble for Cys was also recently developed by Kim et al based on the analyte competing for a metal receptor with a chromogenic indicator.…”

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confidence: 99%

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A Spiropyran‐Based Ensemble for Visual Recognition and Quantification of Cysteine and Homocysteine at Physiological Levels

et al. 2006

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confidence: 99%

“…[10] The approach showed excellent selectivity for Cys over other amino acids including Hcy. Although many reactions and various techniques have been developed for assaying Cys and Hcy, [7][8][9][10] there is still plenty of room for improvement in term of selectivity, sensitivity, and performance with a new interaction mechanism.…”

mentioning

confidence: 99%

A Spiropyran‐Based Ensemble for Visual Recognition and Quantification of Cysteine and Homocysteine at Physiological Levels

et al. 2006

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“…Butyrylthiocholine (BTC) iodide, acetylthiocholine (ATC), and 5,5Ј-dithiobis(2-nitrobenzoic acid) (DTNB) were purchased from Sigma-Aldrich (St. Louis, MO). BES-Thio (2,4-dinitrobenzenesulfonyl fluorescein) was synthesized following procedures described previously (Maeda et al, 2005). Goat anti-rabbit antibodies conjugated with horseradish peroxidase (HRP) enzyme and Supersignal West Pico Chemiluminescent substrate were purchased from Thermo Fisher Scientific (Waltham, MA).…”

Section: Methodsmentioning

confidence: 99%

Identification of Human Butyrylcholinesterase Organophosphate-Resistant Variants through a Novel Mammalian Enzyme Functional Screen

Zhang

Chen

Ralph

et al. 2012

J Pharmacol Exp Ther

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Human butyrylcholinesterase (hBChE) is currently being developed as a detoxication enzyme for the catalytic hydrolysis or stoichiometric binding of organophosphates (OPs). Previously, rationally designed hBChE mutants (G117H and E197Q) were reported in the literature and showed the feasibility of engineering OP hydrolytic functional activity into hBChE. However, the OP hydrolysis rate for G117H is too low for clinical utility. Additional OP-resistant hBChE variants with greater hydrolysis rates are needed as OP nerve-agent countermeasures for therapeutic utility. As described herein, a directed molecular evolution process was used to identify amino acid residues that contribute to OP-resistant functional activity of hBChE variants. In this article, we describe the development and validation of a novel method to identify hBChE variants with OP-resistant functional activity (decreased rate of OP inhibition). The method reported herein used an adenoviral protein expression system combined with a functional screening protocol of OP nerve-agent model compounds that have been shown to have functional properties similar to authentic OP nerveagent compounds. The hBChE screening method was robust for transfection efficiency, library diversity, and reproducibility of positive signals. The screening approach not only identified the previously reported hBChE G117H variant, but also identified a series of additional hBChE variants, including hBChE G117N, G117R, E197C, and L125V, that exhibited OP-resistant functional activities not reported previously. The mammalian functional screening approach can serve as a cornerstone for further optimization and screening for OPresistant hBChEs for potential therapeutic applications.

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“…AChE activity and inhibition behaviors are traditionally monitored with absorption spectroscopy using the Ellman's reagent [42]. Fluorometric approaches with good sensitivity have been reported for screening AChE inhibitors [43][44][45]. Chemiluminescent probes are also available for AChE activity assays and inhibitor screening [46].…”

Section: Colorimetric Assay For Ache Activity and Inhibitor-screeningmentioning

confidence: 99%

Colorimetric detection of glucose and an assay for acetylcholinesterase with amine-terminated polydiacetylene vesicles

Xue

Zhang

et al. 2011

Chin. Sci. Bull.

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The colorimetric response of amine-terminated polydiacetylene (PDA) vesicles was initially demonstrated by varying the pH of the solution. Convenient colorimetric methods to detect glucose and acetylcholinesterase (AChE) activity were successfully established using amine-terminated PDA vesicles by taking advantage of the following features: (1) the amine-terminated PDA vesicles undergo a colorimetric transition as the pH of the solution changes; (2) glucose can be oxidized to gluconic acid in the presence of glucose oxidase; and (3) AChE catalyzes the hydrolysis of acetylcholine to acetic acid. The visual detection of glucose levels and AChE activity showed good selectivity and acceptable sensitivity. The detection limit of glucose was ~2.5 μmol/L and the level of AChE activity was assayed as low as 10.0 mU/mL. Moreover, the amine-terminated PDA vesicles can be used for screening the activity of inhibitors against AChE. polydiacetylene vesicles, glucose, glucose oxidase, acetylcholinesterase Citation:Xue W X, Zhang D Q, Zhang G X, et al. Colorimetric detection of glucose and an assay for acetylcholinesterase with amine-terminated polydiacetylene vesicles.

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2,4‐Dinitrobenzenesulfonyl Fluoresceins as Fluorescent Alternatives to Ellman's Reagent in Thiol‐Quantification Enzyme Assays

Cited by 88 publications

References 23 publications

A Spiropyran‐Based Ensemble for Visual Recognition and Quantification of Cysteine and Homocysteine at Physiological Levels

A Spiropyran‐Based Ensemble for Visual Recognition and Quantification of Cysteine and Homocysteine at Physiological Levels

Identification of Human Butyrylcholinesterase Organophosphate-Resistant Variants through a Novel Mammalian Enzyme Functional Screen

Colorimetric detection of glucose and an assay for acetylcholinesterase with amine-terminated polydiacetylene vesicles

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