title: A fluorimetric assay for human reticulocyte 15-lipoxygenase-1 activity.
abstract: A rapid and sensitive fluorescence-based assay for the determination of human 15-lipoxygenase-1 (15-LOX-1) activity is described in this article. The assay utilizes the ability of 15-LOX-1-generated lipid hydroperoxides to oxidize nonfluorescent dihydrorhodamine 123, producing the highly fluorescent dye rhodamine 123. Formation of rhodamine 123 can be monitored through fluorescence spectroscopy using Ex/Em of 500 nm/536 nm. The IC(50) values of three well-known 15-LOX-1 inhibitors, nordihydroguaiaretic acid, quercetin, and fisetin, were evaluated in 96- and 384-well formats, and they conform to previously reported data. We believe this assay can be broadly used for the discovery of novel lipoxygenase inhibitors.
authors.affiliation: Shanghai Key Laboratory of New Drug Design, Shanghai Key Laboratory of Chemical Biology, School of Pharmacy, East China University of Science and Technology, Shanghai 200237, China.
authors.forename: Weiqiang
authors.initials: W
authors.lastname: Lu
authors.forename: Xue
authors.initials: X
authors.lastname: Zhao
authors.forename: Shien
authors.initials: S
authors.lastname: Zou
authors.forename: Jin
authors.initials: J
authors.lastname: Huang
mesh_terms: D001093:Arachidonate 15-Lipoxygenase / Q000032:analysis* / Q000235:genetics / Q000378:metabolism; D005470:Fluorometry; D057809:HEK293 Cells; D006801:Humans; D016859:Lipoxygenase Inhibitors / Q000737:chemistry; D011794:Quercetin / Q000737:chemistry; D012156:Reticulocytes / Q000201:enzymology*; D012235:Rhodamines / Q000737:chemistry; D014162:Transfection
chemical_list: D016859:Lipoxygenase Inhibitors; D012235:Rhodamines; C058319:dihydrorhodamine 123; D011794:Quercetin; D001093:Arachidonate 15-Lipoxygenase