蛋白质、生物蛋白中蛋白质分子量检测方案(毛细管电泳仪)

HIGH PERFORMANCECAPILLARYELECTROPHORESIS SYSTEMCAPELseries DETERMINATION OF PROTEINS CONCENTRATIONS AND THEIR MOLECULAR WEIGHTS BY CAPILLARY GEL ELECTROPHORESIS INTRODUCTION The method enables fast separation of proteins according to their molecular weights (Mw) with subsequentprotein quantification in protein-containing samples. MEASUREMENT METHOD Capillary electrophoresis for the separation and determination of proteins is based on the differentialmigration of SDS-protein complexes in a narrow fused-silica capillary filled with a low viscous gel， under theinfluence of the applied electric field. Detection of proteins is performed based on their own absorbance at220 nm. Due to the presence of a low viscous gel SDS-protein complexes are separated only according totheir Mw and thus it enables also to determine Mw of an unknown protein(s). ADVANTAGES OF THE CAPILLARY ELECTROPHORESIS METHOD Compared with protein separation with SDS-PAGE， capillary electrophoresiss |has several importantadvantages： Full automatisation ● Direct protein quantification ● Absence of coloring Low analysis cost Short analysis time EQUIPMENTAND REAGENTS The"CAPEL-105/105M”capillary electrophoresis system is used in measurements. Data acquisition， collection， processing and output are performed using a personal computer running under“WINDOWS@2000/XP"operating system with installed dedicated software package for acquisition andprocessing of chromatography data. All reagents must be of analytical grade or higher. Sample preparation， capillary conditioning and analysis are done according to the detailed protocol， includedin the kit. EXAMPLE OF A REAL ANALYSIS Buffer： for SDS-protein complex analysis Capillary： Leff/ Ltot 31/40 cm； ID 75 um Injection： 15 kV for 15 sec Voltage： -25 kV Temperature： +25°C Detection： 220 nmSample： standard proteins with different Mw (from 14.4 kDa to 116 kDa) 17 mAU DEPENDENCE OF LOG Mw OF PROTEINS FROM THEIR MIGRATION TIME This dependence enables direct determination of molecular weight of the unknown protein(s) based on itsmigration time. The contents on this paper are subject to change without notice. WWW.LUMEX.BIZ 毛细管电泳法能够根据蛋白质的分子量快速分离蛋白质并在含有蛋白质的样品中进行蛋白质定量。该法是基于SDS蛋白复合物在一个狭窄的熔融石英毛细管填充低粘度凝胶，在施加电场的作用下进行差分迁移，对其在220 nm波长处的吸光度进行检测。由于低粘度凝胶的存在，SDS蛋白复合物仅根据它们的分子量大小进行分离，因此它也能够确定未知蛋白质的分子量。与SDS-PAGE分离蛋白质相比，毛细管电泳具有几个重要的优点：1.完全自动化；2.直接蛋白质定量；3.无需染色；4.低分析成本；5.分析时间短。