Harvey M D, Bandilla D, Banks P R
Department of Chemistry and Biochemistry, Concordia University, Montreal, QC, Canada.
Electrophoresis. 1998 Sep;19(12):2169-74. doi: 10.1002/elps.1150191221.
Picomolar limits of detection are obtained using the noncovalent, fluorogenic dye, Sypro Red. The size separation of four commonly used sodium dodecyl sulfate-capillary gel electrophoresis (SDS-CGE) molecular weight markers with 8% linear polyacrylamide (PAA) as the sieving matrix is used to construct a calibration curve for molecular weight determinations. SDS-CGE purity and molecular weight determination of purified chorismate mutase-prephenate dehydrogenase (CMPD) from Escherichia coli is shown to be comparable in accuracy with slab gel SDS-polyacrylamide gel electrophoresis (SDS-PAGE). A migration time precision study indicates excellent reproducibility. Sypro red labeling of SDS-bovine serum albumin (SDS-BSA) complexes at nanomolar protein concentrations suggests assay detection limits surpassing those of silver staining. This detectability exceeds that achieved in previous SDS-CGE laser-induced fluorescence studies. This approach is expected to be easily adapted for use with commercial polymer formulations and automated instrumentation.
使用非共价荧光染料Sypro Red可实现皮摩尔级别的检测限。以8%线性聚丙烯酰胺（PAA）作为筛分基质，对四种常用的十二烷基硫酸钠-毛细管凝胶电泳（SDS-CGE）分子量标准品进行尺寸分离，用于构建分子量测定的校准曲线。结果表明，采用SDS-CGE对来自大肠杆菌的纯化分支酸变位酶-预苯酸脱氢酶（CMPD）进行纯度分析和分子量测定，其准确性与平板凝胶SDS-聚丙烯酰胺凝胶电泳（SDS-PAGE）相当。迁移时间精密度研究表明具有出色的重现性。在纳摩尔蛋白质浓度下对SDS-牛血清白蛋白（SDS-BSA）复合物进行Sypro红标记，表明检测限超过银染法。这种可检测性超过了以往SDS-CGE激光诱导荧光研究的水平。预计这种方法可轻松适用于商业聚合物配方和自动化仪器。
