Law Wai S, Zhao Jian H, Li Sam F Y
Department of Chemistry, National University of Singapore, Singapore, Republic of Singapore.
Electrophoresis. 2005 Sep;26(18):3486-94. doi: 10.1002/elps.200500255.
Field-amplified sample stacking (FASS) is used to separate basic proteins in a poly-(vinyl alcohol)-coated bubble cell capillary. To our knowledge, this is the first paper describing the on-column stacking of proteins (as cations) using FASS in bubble cell capillary. The bubble cell capillary is fabricated using a one-step method. Cetyltrimethylammonium chloride is added into the running buffer to reverse the EOF and, thus, to pump the water plug out during the sample stacking step. The effect of the water plug lengths and sample injection durations were investigated and optimized. The results obtained were compared with those for the normal capillary without bubble cell in terms of resolution and sensitivity enhancement. Under the optimal condition, this method can improve the sensitivity of the peak areas ranging from 5000- to 26 000-fold. The RSDs (n = 5) of the migration time and peak area are satisfactory (less than 0.6 and 12%, respectively). Application of the capillary electrophoresis method with bubble cell, FASS, and UV detection thereby leads to the determination of these proteins at concentrations ranging from 3 to 10 ng/mL, based on a signal-to-noise ratio of 3:1.
场放大进样堆积(FASS)用于在聚乙烯醇涂层气泡池毛细管中分离碱性蛋白质。据我们所知,这是第一篇描述在气泡池毛细管中使用FASS对蛋白质(作为阳离子)进行柱上堆积的论文。气泡池毛细管采用一步法制备。将十六烷基三甲基氯化铵添加到运行缓冲液中以反转电渗流,从而在进样堆积步骤中将水塞泵出。研究并优化了水塞长度和进样持续时间的影响。将获得的结果与没有气泡池的普通毛细管在分离度和灵敏度提高方面的结果进行了比较。在最佳条件下,该方法可将峰面积的灵敏度提高5000至26000倍。迁移时间和峰面积的相对标准偏差(n = 5)令人满意(分别小于0.6%和12%)。因此,采用带有气泡池、FASS和紫外检测的毛细管电泳方法,基于3:1的信噪比,可测定浓度范围为3至10 ng/mL的这些蛋白质。