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首页> 外文期刊>Journal of immunoassay >A simple method of electroelution of individual protein bands from SDS polyacrylamide gels for direct study in cellular assays.
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A simple method of electroelution of individual protein bands from SDS polyacrylamide gels for direct study in cellular assays.

机译：一种从SDS聚丙烯酰胺凝胶中电洗脱单个蛋白质条带的简单方法，可直接在细胞测定中进行研究。

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摘要

A very simple and effective procedure which allows simultaneous electroelution of separated proteins from SDS polyacrylamide gel into small quantity of elution buffer is described. Elution parameters have been optimized for maximum possible recovery (50-60%). Protein fractions were collected in physiological buffer and an efficient removal of SDS have been obtained, thus fractions collected were suited for direct testing in cell cultures. Method was used to investigate human T-cell responses to purified secreted M tuberculosis H37Rv proteins. Eight low molecular weight (M.w. range 10 kD to 25 kD) culture filtrate proteins were purified in quantities, sufficient for immunological characterization. Lymphocyte proliferative responses and cytokine release pattern from tuberculosis patients, healthy contacts and healthy controls were studied on stimulation with purified culture filtrate proteins. Immunologically important M.tuberculosis proteins were identified by using this method. This approach should be applicable to the rapid identification and characterization of any interesting T cell antigen.
机译：描述了一种非常简单有效的方法，该方法可将分离的蛋白质从SDS聚丙烯酰胺凝胶中同时洗脱到少量洗脱缓冲液中。洗脱参数已经过优化，可实现最大回收率（50-60％）。将蛋白质级分收集在生理缓冲液中，并且可以有效去除SDS，因此收集的级分适合在细胞培养物中直接检测。方法用于研究人T细胞对纯化的分泌性M结核H37Rv蛋白的反应。大量纯化了八种低分子量（分子量范围从10 kD到25 kD）的培养滤液蛋白，足以进行免疫学表征。研究了纯化培养滤液蛋白对结核病患者，健康人和健康对照者淋巴细胞增殖反应和细胞因子释放模式的影响。使用此方法鉴定了具有免疫学意义的结核分枝杆菌蛋白。这种方法应适用于任何有趣的T细胞抗原的快速鉴定和表征。

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