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

✍ Scribed by Hatsuo Maeda; Hiromi Matsuno; Mai Ushida; Kohei Katayama; Kanako Saeki; Norio Itoh

• Book ID: 101529938
• Publisher: John Wiley and Sons
• Year: 2005
• Tongue: English
• Weight: 388 KB
• Volume: 117
• Category: Article
• ISSN: 0044-8249
• DOI: 10.1002/ange.200500114

✦ Synopsis

Since its first description in 1959, [1] 5,5'-dithio-bis(2-nitrobenzoic acid) (DTNB or Ellmans reagent) has been used widely for the quantification of thiols, especially for assays of various enzymes including acetyl-and butyrylcholinesterase (AChE and BChE, respectively) [2] in which substrates release thiols through enzymatic reactions. [3] 4,4'-Bipyridyl disulfide [4] and 5-(2-aminoethyl)dithio-2-nitrobenzoic acid [3] were also shown to be useful for colorimetric analyses of thiols. However, fluorescent or chemiluminescent thiol probes would be more practical than these chromogenic probes for enzyme assays, especially for high-throughput programs to screen enzyme inhibitors as new drugs. Recently, the use of a dioxetane-disulfide derivative as a chemiluminescent probe in place of DTNB led to an improved sensitivity of the AChE assay with acetylthiocholine (ASCh) by a factor of ten. [5] However, the assay required a rather tedious procedure in