One-dimensional protein analysis of an HT29 human colon adenocarcinoma cell
One-dimensional protein analysis of an HT29 human colon adenocarcinoma cell
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DOI：
10.1021/ac990694y
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发表时间：
2000-01-15
影响因子：
7.4
通讯作者：
Dovichi, NJ
中科院分区：
文献类型：
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作者：
Zhang, ZR;Krylov, S;Dovichi, NJ
将单个HT29人结肠腺癌细胞引入熔融石英毛细管裂解，用荧光试剂3-(2-呋喃甲酰)喹啉-2-甲醛荧光标记蛋白质含量。标记的蛋白质在胶束下缓冲液中通过毛细管电泳法分离，并在柱后鞘-NOW试管中用激光诱导荧光检测。解决了数十个组件。为了证实这些成分是蛋白质，进行了大量的实验。单细胞电泳图的大多数成分与从细胞培养中制备的蛋白质提取物的30-100 kDa部分中的成分具有相同的迁移率。通过将样品与从SDS-PAGE凝胶中获得的纯化蛋白共同注射，其中一个组分被鉴定为类似100 kDa的蛋白质。不同细胞的蛋白质表达差异很大，但平均表达与从10(6)个细胞制备的蛋白质提取液观察到的一致。
A single HT29 human colon adenocarcinoma cell was introduced into a fused-silica capillary and lysed, and the protein content was fluorescently labeled with the fluorogenic reagent 3-(2-furoyl)quinoline-2-carboxaldehyde. The labeled proteins were separated by capillary electrophoresis in a submicellar buffer and detected by laser-induced fluorescence in a postcolumn sheath-now cuvette. Several dozen components were resolved. A number of experiments were done to verify that these components were proteins. Most components of the single-cell electropherogram had the same mobility as components present in the 30-100 kDa fraction of a protein extract prepared from the cell culture. One component was identified as a similar to 100 kDa protein by co-injecting the sample with purified protein obtained from an SDS-PAGE gel. Protein expression varied significantly between cells, but the average expression was consistent with that observed from a protein extract prepared from 10(6) cells.