Isoluminol-enhanced chemiluminescence: A sensitive method to study the release of superoxide anion from human neutrophils
Isoluminol-enhanced chemiluminescence: A sensitive method to study the release of superoxide anion from human neutrophils
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DOI：
10.1016/0891-5849(95)02189-2
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发表时间：
1996-01-01
影响因子：
7.4
通讯作者：
Dahlgren, C
中科院分区：
文献类型：
--
作者：
Lundqvist, H;Dahlgren, C
建立了一种测定人中性粒细胞呼吸爆发产物的化学发光法。尽管CL反应是过氧化物酶依赖性的，但发现过氧化氢不参与光产生反应。将吞噬细胞与异鲁米诺混合，异鲁米诺是一种化学发光底物，仅检测细胞外释放的氧物质。由于释放的细胞髓过氧化物酶的可用性是反应中的限制因素，因此必须向测量系统中添加额外的过氧化物酶（辣根过氧化物酶; HRP）。从超氧化物歧化酶（SOD）几乎完全抑制佛波醇肉豆蔻酸酯（PMA）和甲酰-甲硫氨酰-亮氨酰-苯丙氨酸（fMLP）的反应这一事实，我们得出结论，O-2（.-）是测量的反应氧物质。所描述的测定系统非常适合于从活化的中性粒细胞释放超氧阴离子的实时研究。利用该技术，释放O-2（.-）可以从250个中性粒细胞中检测到。
A very sensitive chemiluminescence (CL) method was developed for the determination of respiratory burst products generated by the NADPH-oxidase in human neutrophils. Despite the fact that the CL reaction is peroxidase dependent, hydrogen peroxide was found not to participate in the light generating reaction. Phagocytic cells were mixed with isoluminol, a chemiluminescence substrate that detects extracellularly released oxygen species only, Owing to the fact that the availability of released cellular myeloperoxidase is a limiting factor in the reaction, extra peroxidase (horseradish peroxidase; HRP) has to be added to the measuring system. From the fact that the response to phorbol myristate acetate (PMA), as well as to formyl-methionyl-leucyl-phenylalanine (fMLP) was almost totally inhibited by superoxide dismutase (SOD), we conclude that O-2(.-) is the reacting oxygen species measured. The assay system described is well suited for real-time studies of superoxide anion release from activated neutrophils. With the technique, the release of O-2(.-) can be detected from as few as 250 neutrophils.