Wang S, Rosengren L E, Karlsson J E, Stigbrand T, Haglid K G
Institute of Neurobiology, University of Göteborg, Sweden.
J Neurosci Methods. 1990 Aug;33(2-3):219-27. doi: 10.1016/0165-0270(90)90025-b.
Immunoassays for quantitative determinations of the S-100 protein, the glial fibrillary acidic protein, the neuron specific enolase and the neurofilament proteins with molecular weight of 68 and 200 kDa in hot SDS sonicated rat brain extracts have been developed and characterized. The assays utilize a dot immunobinding technique, poly- or monoclonal antibodies and 125I-protein A. The SDS-sonication procedure was not found to affect the radioactivity recovery in the assay of the soluble S-100 protein or the neuron specific enolase. All 5 antigens can be measured with a within-assay variance below 10%. Even at a coefficient of variation less than or equal to 5%, the working ranges are approximately 30-100-fold with regard to the different antigens. It was found that gelatin-coated nitrocellulose membranes considerably increase the recovered radioactivity in the assay of the purified bovine S-100 protein, possibly by protein-protein interaction. This effect was not observed when SDS-sonicated rat brain extracts were assayed. The assay appears to be reproducible, convenient and rapid, and provides a high degree of precision in the determination of large number of samples.
已开发并表征了用于定量测定热SDS超声处理的大鼠脑提取物中S-100蛋白、胶质纤维酸性蛋白、神经元特异性烯醇化酶以及分子量为68和200 kDa的神经丝蛋白的免疫测定方法。这些测定采用斑点免疫结合技术、多克隆或单克隆抗体以及125I-蛋白A。在可溶性S-100蛋白或神经元特异性烯醇化酶的测定中，未发现SDS超声处理程序会影响放射性回收率。所有5种抗原的测定批内变异均低于10%。即使变异系数小于或等于5%，不同抗原的工作范围也约为30至100倍。发现明胶包被的硝酸纤维素膜在纯化牛S-100蛋白的测定中可显著提高回收的放射性，这可能是通过蛋白质-蛋白质相互作用实现的。在测定SDS超声处理的大鼠脑提取物时未观察到这种效应。该测定方法具有可重复性、方便快捷，并且在大量样品的测定中提供了高度的精确性。
