Bromobimane probes for thiols

Research output: Contribution to journalComment/debate

Abstract

This chapter discusses bromobimane probes for thiols and describes the use of four bromobimanes for fluorescent labeling of biochemical and biological systems. The four bromobimanes are (1) mBBr, (2) bBBr, (3) qBBr, and (4) SBBr. The bromobimanes are essentially nonfluorescent and are relatively stable in the dry state when stored in the dark. Chromatography on thin-layer silica yields a yellow nonfluorescent spot that develops a blue fluorescence after several minutes of exposure to 360-nm light. The change is a convenient characteristic for the identification of the bromobimanes and suggests sensitivity to light (photolysis). Bromobimanes in solution react with small thiols (e.g., the tripeptide thiol glutathione [GSH]), and with reactive protein thiol groups (e.g., hemoglobin). The reactions of bromobimanes with thiols are second order and dependent on pH, the active nucleophile being the thiolate anion, such as GS. The reaction of bromobimane with a thiolate converts the nonfluorescent agent into water-soluble fluorescent products. The chapter discusses the chemical and photophysical properties of bromobimanes, labeling procedures, labeling of thiols in tissues, and other applications of thiol labeling.

Original languageEnglish
Pages (from-to)133-148
Number of pages16
JournalMethods in Enzymology
Volume251
Issue numberC
DOIs
StatePublished - 1 Jan 1995

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