title: An enzymatic colorimetric assay for glucose-6-phosphate.
abstract: A specific colorimetric assay for the determination of glucose-6-phosphate (G6P) was developed. This assay is based on the oxidation of G6P in the presence of glucose-6-phosphate dehydrogenase (G6PD) and nicotinamide adenine dinucleotide phosphate (NADP(+)); the NADPH thereby generated reduces the tetrazolium salt WST-1 [2-(4-indophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H tetrazolium, monosodium salt] to water-soluble yellow-colored formazan with 1-methoxy-5-methylphenazium methylsulfate (1-mPMS) as an electron carrier. The assay is optimized for reaction buffer pH, enzyme/dye concentration, and reaction time course. The limit of detection of the assay is 0.15 μM (15 pmol/well). The usefulness of the assay is demonstrated by the accurate measurement of the G6P concentration in fetal bovine serum (FBS).
authors.affiliation: Department of Ophthalmology and Visual Sciences, University of Michigan Medical School, Ann Arbor, MI 48105, USA.
authors.forename: Aiping
authors.initials: A
authors.lastname: Zhu
authors.forename: Roberto
authors.initials: R
authors.lastname: Romero
authors.forename: Howard R
authors.initials: HR
authors.lastname: Petty
mesh_terms: D000818:Animals; D002138:Calibration; D002417:Cattle; D003124:Colorimetry / Q000379:methods*; D057075:Enzyme Assays / Q000379:methods*; D019298:Glucose-6-Phosphate / Q000737:chemistry / Q000378:metabolism*; D005954:Glucosephosphate Dehydrogenase / Q000378:metabolism*; D006358:Hot Temperature; D007934:Leuconostoc / Q000201:enzymology; D015203:Reproducibility of Results; D044967:Serum
chemical_list: D019298:Glucose-6-Phosphate; D005954:Glucosephosphate Dehydrogenase