Kit Size | 500+300 |
---|
Myeloperoxidase (MPO) is a highly cationic glycosolated hemoprotein that has a molecular weight of 144kD. The hemoprotein consists of two dimers linked via a disulfide bridge. Each dimmer is composed of a heavy (53kD) and light (15kD) subunit. Each heavy chain contains an independently acting protoporphyrin group containing a central iron (1-5). MPO is present in the azurophilic granules of polymorphonuclear leukocytes (PMNs) and is unique to neutrophils and monocytes. However, monocytes contain only one third of the MPO found in PMN’s. MPO utilizes H202 produced by the neutrophils to oxidize a varity of aromatic compounds to give substrate radicals for bactericidal activity (4 review). This enzyme is unique however in that it can oxidize chloride ions to produce a strong nonradical oxidant,HOCl. HOCl is the most powerful bactericidal produced by neutrophils (4 review). Excessive production of these radicals can cause oxidative stress leading to oxidative tissue injury.
Chlorination Reaction:
H2O2 + MPO + Cl- —-> HOCl + APF (non fluorescent) —-> Fluorescent Dye
Excitation:488nm; Emission: 515-530nm
Peroxidation Reaction:
H2O2 + Detection reagent (non-fluorescent)+ MPO —–> fluorescent analog
Excitation 530-571nm Emission 590-600nm
Figure 1. In this figure, the MPO standard curve was serially diluted in 1X Reaction Buffer. Reaction cocktail (RC) was prepared as described (without EPO inhibitor). Next 50µL of MPO standard and 50µL of RC was added to individual well of 96- well black plates. The plate was incubated at room and temperature in the dark. Data collected Ex:530nm, Em:590nm
Figure 2. Red Blood Cell AChE (RBC-AChE) was purified and protein concentration determined using the BCA Protein Assay Kit (Pierce). The RBC-AChE was titrated in 1X reaction buffer and activity determined using the Fluoro: AChE kit. Acetylcholine concentration = 1mM final. In the graph the background value has been subtracted (0 RBC-AChE) to generate standard curve.
Document Title |
MPOHOCL Protocol |
Fluoro MPOHOCL Datasheet |
msds.Mpohocl |
Reference |
Olsen, R. L. & Little, c. (1983) Biochem. J. 209, 781-787. |
Nauseef, W. M., and Malech, H. L. (1986) Blood, 67, 1504-1507. |
Andrews, P. C., Parnes, C., and Krinsky, N. I. (1984) Arch. Biochem. Biophys., 228, 439-442. |
Mark B. Hampton, Anthony J. Kettle, and Christine C. Winterbourn. Inside the Neutrophil Phagosome: Oxidants, Myeloperoxidase, and Bacterial Killing. Blood, Vol. 92 No. 9 (November 1), 1998: pp. 3007-3017 |
Mark B. Hampton, Anthony J. Kettle, and Christine C. Winterbourn. Inside the Neutrophil Phagosome: Oxidants, Myeloperoxidase, and Bacterial Killing. Blood, Vol. 92 No. 9 (November 1), 1998: pp. 3007-3017 |
Andrews, P.C., Parnes, C. & Krinsky, N.I. (1984) Comparison of myeloperoxidase and hemi-myeloperoxidase with respect to catalysis, regulation, and bacterial activity. Arch. Biochem. Biophys. 228, 439–442. |
Bolognesi ML et al. Propidium-based polyamine ligands as potent inhibitors of acetylcholinesterase and acetylcholinesterase-induced amyloid-beta aggregation. J Med Chem. 2005 Jan 13;48(1):24-7. |
Ken-ichi Setsukinai, Yasuteru Urano, Katsuko Kakinuma, Hideyuki J. Majima , and Tetsuo Nagano. Development of Novel Fluorescence Probes That Can Reliably Detect Reactive Oxygen Species and Distinguish Specific Species. THE JOURNAL OF BIOLOGICAL CHEMISTRY Vol. 278, No. 5, Issue of January 31, pp. 3170–3175, 2003 |
Part# | Reagent | Temperature |
Part# 4007 | Detection Reagent, 1 Vial | -20C |
Part# 3002 | 10X Assay Buffer, 60mL | 2-8C |
Part# 3012 | Hydrogen Peroxide, 1000µL of a Stabilized 3% Solution | 2-8C |
Part# 6015 | Myeloperoxidase, 1 Vial at 30Units/mL | 2-8C |
Part# 4011 | APF, 1 Vial | 2-8C |
Cell Signaling Technology (CST) 是一家由科学家创立的私营家族公司,致力于提供全球最高品质的创新研究和诊断产品,加速生物学认知以及实现个体化医疗。CST坚持自主生产和严格验证,其高质量的产品和专业的研发精神已被全球客户认可,被公认/票选为细胞信号研究的金标准、最佳抗体*、研究者的选择*、PTM(蛋白翻译后修饰)年度抗体公司*、十年抗体品牌*等(*来自CiteAb、LISA数据报告)。
CST提供最高品质的特色信号蛋白及磷酸化、甲基化、乙酰化、泛素化、SUMO化等翻译后修饰抗体,蛋白翻译后修饰筛选试剂盒及服务,还有偶联抗体,二抗,ChIP试剂盒,细胞因子,ELISA试剂盒,细胞检测试剂盒,蛋白实验配套试剂等,为您提供蛋白相关实验的一站式解决方案。