Improved Separation of Microheterogeneities and Isoforms of Proteins by Capillary Electrophoresis Using Segmental Filling with SDS and PEO in the Background Electrolyte
Resource
Analytical Chemistry 74 (18): 4828-4834
Journal
Analytical Chemistry
Journal Volume
74
Journal Issue
18
Pages
4828-4834
Date Issued
2002
Date
2002
Author(s)
Abstract
To improve the separation efficiency while achieving high sensitivity for the analysis of proteins' microheterogeneity, a segmental-filling technique has been developed and tested in capillary electrophoresis with laser-induced native fluorescence using a pulsed Nd:YAG laser. Using a short plug of SDS applied to the capillary and the anticonvectant poly(ethylene oxide) (PEO), the microheterogeneities of a number of proteins with pI values ranging from 4.5 to 11.1 could be detected. This high resolving power is due to reduced adsorption on the capillary wall, sieving, and the interaction with SDS. Consequently, the length and the concentration of the SDS plug play a significant role in determining the resolution and sensitivity. The method has been applied to the analysis of salivary and cerebrospinal fluid (CSF) samples. Without any sample pretreatment, using a 10-s 1x SDS plug, six alpha-amylase isoforms in a salivary sample were resolved in 17 min and three more peaks were detected in a CSF sample. With simplicity, high resolving power, and rapidity, the method has shown great potential for proteomics.
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Type
journal article
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