Design of a Practical Fluorescent Probe for Superoxide Based on Protection–Deprotection Chemistry of Fluoresceins with Benzenesulfonyl Protecting Groups

書誌事項

公開日
2007-02-08
権利情報
  • http://onlinelibrary.wiley.com/termsAndConditions#vor
DOI
  • 10.1002/chem.200600522
公開者
Wiley

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説明

<jats:title>Abstract</jats:title><jats:p>A strategy for designing probes based on protection–deprotection chemistry involving fluoresceins and their benzenesulfonyl (BES) derivatives has led to the development of a much more practical superoxide (O<jats:sub>2</jats:sub><jats:sup>−.</jats:sup>) probe than the previously reported bis(2,4‐dinitro‐BES) tetrafluorofluorescein (<jats:bold>6 a</jats:bold>). Examination of various BES derivatives, developed from the starting point of the prototype probe <jats:bold>6 a</jats:bold>, yielded 4,5‐dimethoxy‐2‐nitro‐BES tetrafluorofluorescein (BESSo; <jats:bold>7 j</jats:bold>) as the optimal reagent. A microtiter plate assay with BESSo showed a tenfold improved detection limit for O<jats:sub>2</jats:sub><jats:sup>−.</jats:sup> compared with such an assay based on <jats:bold>6 a</jats:bold>. BESSo showed markedly better specificity for O<jats:sub>2</jats:sub><jats:sup>−.</jats:sup> than for GSH or other reactive oxygen species, and this specificity was significantly higher than that of Fe<jats:sup>2+</jats:sup> and some reducing enzymes. These features have resulted in the development of an assay based on BESSo that is capable of providing more unambiguous results for O<jats:sub>2</jats:sub><jats:sup>−.</jats:sup> release from neutrophils, with or without stimulation by phorbol myristate acetate, as compared with an assay based on <jats:bold>6 a</jats:bold>. Intracellular generation of O<jats:sub>2</jats:sub><jats:sup>−.</jats:sup> in human Jurkat T cells stimulated by butyric acid has been measured by using flow cytometry and fluorescence microscopy utilizing the acetoxymethyl derivative of BESSo.</jats:p>

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