Immunological detection and quantification of oxidized proteins by labelling with digoxigenin
Immunological detection and quantification of oxidized proteins by labelling with digoxigenin
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DOI：
10.1271/bbb.62.419
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发表时间：
1998-03-01
影响因子：
1.6
通讯作者：
Mateos-Nevado, MD
中科院分区：
文献类型：
--
作者：
Bautista, J;Mateos-Nevado, MD
开发了一种基于地高辛/抗地高辛系统的免疫测定法，用于检测和定量蛋白质氧化损伤产生的羰基部分。牛血清白蛋白 (BSA) 被抗坏血酸/Fe(III)/O-2 组成的羟基自由基生成系统氧化。所得白蛋白衍生的羰基用地高辛酰肼标记，并用与碱性磷酸酶缀合的抗地高辛抗体通过点印迹进行检测。通过光密度分析进行定量。该系统可以检测印迹上皮摩尔量的羰基。该检测的灵敏度范围为 1.26 至 126 pmole。该方法的另一个特点是将其应用于复杂的蛋白质混合物（匀浆）来分析单个蛋白质的氧化状态，如大鼠肠刷状缘膜匀浆所示。
An immunological assay, based on the digoxigenin/antidigoxigenin system, was developed to detect and quantify carbonyl moieties that result from oxidative damage to proteins. Bovine serum albumin (BSA) was oxidized by a hydroxyl radical-generating system consisting of ascorbate/Fe(III)/O-2. The resulting albumin-derived carbonyls were labelled with digoxigenin-hydrazide and detected by dot blotting with an anti-digoxigenin antibody conjugated to alkaline phosphatase. Quantification was Carried out by a densitometric analysi. This system allows the detection of a pmole-amount of carbonyl groups on blots. The assay covers a range of sensitivity from 1.26 to 126 pmoles. Another feature of this method is its application to a complex protein mixture (homogenate) to analyze the oxidative status of individual proteins, as are shown for intestinal brush border membrane homogenate of a rat.