A fluorimetric assay for human reticulocyte 15-lipoxygenase-1 activity

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• 作者：Weiqiang Lu^a ; Xue Zhao^a ; Shien Zou^b ; Jin Huang^a ; ^{huangjin@ecust.edu.cn}
• 关键词：15-Lipoxygenase-1 ; Inhibitor ; Fluorimetric assay ; Dihydrorhodamine 123
• 刊名：Analytical Biochemistry
• 出版年：2012
• 期刊代码：93_00032697
• 类别：imm
• 出版时间：1 July, 2012
• 卷：426
• 期：1
• 页码：66-68
• 文件大小：272 K

摘要

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 IC₅₀ 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.