Skip Navigation
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Your Environment. Your Health.

Publication Detail

Title: Fluorescence-based microtiter plate assay for glutamate-cysteine ligase activity.

Authors: White, Collin C; Viernes, Hannah; Krejsa, Cecile M; Botta, Dianne; Kavanagh, Terrance J

Published In Anal Biochem, (2003 Jul 15)

Abstract: Glutamate-cysteine ligase (GCL; also known as gamma-glutamylcysteine synthetase) is the rate-limiting enzyme in glutathione (GSH) synthesis. Traditional assays for the activity of this enzyme are based either on coupled reactions with other enzymes or on high-performance liquid chromatography (HPLC) assessment of gamma-glutamylcysteine (gamma-GC) product formation. We took advantage of the reaction of naphthalene dicarboxaldehyde (NDA) with GSH or gamma-GC to form cyclized products that are highly fluorescent. Hepa-1 cells which were designed to overexpress mouse GCL and mouse liver homogenates were used to evaluate and compare the utility of the NDA method with an assay based on monobromobimane derivatization and HPLC analysis with fluorescence detection. Excellent agreement was found between GCL activities measured by HPLC and NDA-microtiter plate analyses. This assay should be useful for high-throughput GCL activity analyses.

PubMed ID: 12814619 Exiting the NIEHS site

MeSH Terms: Animals; Cells, Cultured; Fluorescence; Glutamate-Cysteine Ligase/analysis*; Glutamate-Cysteine Ligase/metabolism*; Hydrogen-Ion Concentration; Liver Extracts; Liver/cytology; Liver/enzymology; Mice; Microchemistry/methods; Time Factors

Back
to Top