Protein carbonyl measurement by a sensitive ELISA method
Protein carbonyl measurement by a sensitive ELISA method
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DOI：
10.1016/s0891-5849(97)00104-4
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发表时间：
1997-01-01
影响因子：
7.4
通讯作者：
Winterbourn, CC
中科院分区：
文献类型：
--
作者：
Buss, H;Chan, TP;Winterbourn, CC
我们描述了一种新的免疫测定蛋白质羰基作为氧化损伤的指标。将蛋白质样品与二硝基苯肼反应，然后吸附到ELISA板的威尔斯孔中，然后用针对蛋白质缀合的二硝基苯肼产生的商业抗体进行探测。然后将生物素偶联的一抗与链霉亲和素-生物素化辣根过氧化物酶反应进行定量。该方法使用氧化白蛋白进行校准，结果与比色羰基测定相关性良好。该方法只需要60 μ g蛋白质，并用于分析血浆和肺吸出物样品中的蛋白质羰基的量。它在0-2.5 nmol/mg蛋白质范围内是敏感的，临床样品在该范围内下降，并在10 nmol/mg蛋白质内呈线性。酶联免疫吸附法测定蛋白质羰基比比色法灵敏度高，具有较好的鉴别能力，值得推广应用。实验和临床样品，特别是在浓度低和只有少量样品可用的情况下。(C)1997年爱思唯尔科学公司
We describe a new immunoassay for measuring protein carbonyls as an index of oxidative injury. Protein samples were reacted with dinitrophenylhydrazine then adsorbed to wells of an ELISA plate before probing with a commercial antibody raised against protein-conjugated dinitrophenylhydrazine. The biotin-conjugated primary antibody was then reacted with streptavidin-biotinylated horseradish peroxidase for quantification. The method was calibrated using oxidized albumin and results correlated well with the colorimetric carbonyl assay. The method required only 60 mu g protein and was used to analyze the amount of protein carbonyls in plasma and lung aspirate samples. It was sensitive in the 0-2.5 nmol/mg protein range within which clinical samples fell and was linear up to 10 nmol/mg protein. The ELISA method for protein carbonyls is more sensitive and discriminatory than the colorimetric assay and should have wide application for analysing. experimental and clinical samples, especially where concentrations are low and where only small amounts of sample are available. (C) 1997 Elsevier Science Inc.