Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript. AbstractThe spectrophotometric/microplate reader assay method for glutathione (GSH) involves oxidation of GSH by the sulfhydryl reagent 5,5鈥?dithio-bis(2-nitrobenzoic acid) (DTNB) to form the yellow derivative 5鈥?thio-2-nitrobenzoic acid (TNB), measurable at 412 nm. The glutathione disulfide (GSSG) formed can be recycled to GSH by glutathione reductase in the presence of NADPH. The assay is composed of two parts: the preparation of cell cytosolic/tissue extracts and the detection of total glutathione (GSH and GSSG). The method is simple, convenient, sensitive and accurate. The lowest detection for GSH and GSSG is 0.103 nM in a 96-well plate. 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Normal alveolar epithelial lining fluid contains high levels of glutathione. Am. J. Physiol. 63, 152鈥?57 (1987).CAS聽 Google Scholar聽 Download referencesAuthor informationAffiliationsDepartment of Environmental Medicine, Lung Biology and Disease Program, University of Rochester Medical Center, Rochester, New York, USAIrfan Rahman聽 聽Aruna KodeDepartment of Biochemistry and Biotechnology, Dr. Ambedkar College, Nagpur, Maharashtra State, IndiaSaibal K BiswasAuthorsIrfan RahmanView author publicationsYou can also search for this author in PubMed聽Google ScholarAruna KodeView author publicationsYou can also search for this author in PubMed聽Google ScholarSaibal K BiswasView author publicationsYou can also search for this author in PubMed聽Google ScholarCorresponding authorCorrespondence to Irfan Rahman.Ethics declarations Competing interests The authors declare no competing financial interests. Rights and permissionsReprints and PermissionsAbout this articleCite this articleRahman, I., Kode, A. Biswas, S. Assay for quantitative determination of glutathione and glutathione disulfide levels using enzymatic recycling method. Nat Protoc 1, 3159鈥?165 (2006). https://doi.org/10.1038/nprot.2006.378Download citationPublished: 31 January 2007Issue Date: December 2006DOI: https://doi.org/10.1038/nprot.2006.378 Victoria K. Mwaeni, James N. Nyariki, Ngalla Jillani, George Omwenga, Mathew Ngugi Alfred Orina Isaac BMC Pharmacology and Toxicology (2021) Heera Ram, Pramod Kumar, Ashok Purohit, Priya Kashyap, Suresh Kumar, Shivani Kumar, Garima Singh, Abdulaziz A. 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