Sheer D G, Yamane D K, Hawke D H, Yuan P M
Applied Biosystems, Foster City, CA 94404.
Biotechniques. 1990 Oct;9(4):486-95.
High-performance electrophoresis chromatography (HPEC) is a recent development that features continuous-elution gel electrophoresis for isolating proteins or peptides in range of 1 to 300 microgram quantities. Column gel electrophoresis is conducted under thermostated conditions, and the field voltage can be varied within a run with a programmable power supply. Applications of this apparatus in protein purification are presented to demonstrate the utility of the (Model 230A) HPEC. These examples include on-line detection with direct analyte recovery of highly purified sample, which mimics high-performance liquid chromatography, for subsequent structure-function characterization. A method to remove salts from sodium dodecyl sulfate electrophoresed samples for subsequent sequencing or amino acid analysis is described. This desalting procedure recovers from 90%-95% of the sample and employs a low molecular weight cut-off membrane during sample centrifugation onto a polyvinylidene difluoride membrane. Subsequent washings are performed to efficiently remove salts, free amino acids and detergents that are known to interfere with sequence analysis. Sequence information such as initial recovery, repetitive yields and chromatogram comparisons are presented to demonstrate the utility of this procedure when used following isolation of sample with HPEC.
高效电泳色谱法（HPEC）是一项最新进展，其特点是采用连续洗脱凝胶电泳法来分离1至300微克量范围内的蛋白质或肽。柱凝胶电泳在恒温条件下进行，场电压可在运行过程中通过可编程电源进行变化。介绍了该仪器在蛋白质纯化中的应用，以证明（230A型）HPEC的实用性。这些例子包括对高度纯化样品进行直接分析物回收的在线检测，这类似于高效液相色谱法，用于后续的结构-功能表征。描述了一种从十二烷基硫酸钠电泳样品中去除盐分以便后续测序或氨基酸分析的方法。该脱盐程序可回收90%-95%的样品，并且在将样品离心到聚偏二氟乙烯膜上的过程中使用低分子量截留膜。随后进行洗涤以有效去除已知会干扰序列分析的盐分、游离氨基酸和去污剂。给出了诸如初始回收率、重复产率和色谱图比较等序列信息，以证明该程序在使用HPEC分离样品后使用时的实用性。
