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Design of a Practical Fluorescent Probe for Superoxide Based on Protection–Deprotection Chemistry of Fluoresceins with Benzenesulfonyl Protecting Groups
Author(s) -
Maeda Hatsuo,
Yamamoto Kayoko,
Kohno Iho,
Hafsi Leila,
Itoh Norio,
Nakagawa Shinsaku,
Kanagawa Naoko,
Suzuki Keiichiro,
Uno Tadayuki
Publication year - 2007
Publication title -
chemistry – a european journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.687
H-Index - 242
eISSN - 1521-3765
pISSN - 0947-6539
DOI - 10.1002/chem.200600522
Subject(s) - chemistry , superoxide , combinatorial chemistry , fluorescence , protecting group , organic chemistry , enzyme , physics , alkyl , quantum mechanics
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 2 −. ) probe than the previously reported bis(2,4‐dinitro‐BES) tetrafluorofluorescein ( 6 a ). Examination of various BES derivatives, developed from the starting point of the prototype probe 6 a , yielded 4,5‐dimethoxy‐2‐nitro‐BES tetrafluorofluorescein (BESSo; 7 j ) as the optimal reagent. A microtiter plate assay with BESSo showed a tenfold improved detection limit for O 2 −. compared with such an assay based on 6 a . BESSo showed markedly better specificity for O 2 −. than for GSH or other reactive oxygen species, and this specificity was significantly higher than that of Fe 2+ 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 2 −. release from neutrophils, with or without stimulation by phorbol myristate acetate, as compared with an assay based on 6 a . Intracellular generation of O 2 −. 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.