Capillary isoelectric focusing-mass spectrometry: analysis of protein mixtures from human body fluids

Biomedical Chromatography : BMC
Nigel J Clarke, Stephen Naylor

Abstract

Isoelectric focusing within a fused silica capillary (cIEF) has proved to be a powerful and practical method for high-resolution separation of analytes from complex biological mixtures. This technique overcomes many of the problems of isoelectric focusing within slab gel media. However current cIEF systems commonly utilize UV detection which limits the detail of analyte structural information that is obtained during analysis. The use of mass spectrometry (MS) as the detection system provides much greater structural information about the detected analytes allowing accurate relative molecular mass (M(r)) determination for proteins and polypeptides. We have constructed a cIEF-MS interface and compared the separation of standard proteins analyzed by cIEF-UV with cIEF-MS. This allowed rapid optimization of the cIEF-MS system performance. Further we have demonstrated the use of MS as a detection system provides accurate M(r) information and can provide analyte modification details. These factors increase the likelihood of absolute identification for physiological proteins within complex in vivo-derived mixtures. To demonstrate the value of cIEF-MS in such analyses we have undertaken an examination of cerebrospinal fluid (CSF), and te...Continue Reading

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Citations

Aug 14, 2012·Proteomics·Shuai Sherry ZhaoDavid D Y Chen
Mar 3, 2005·Journal of Pharmaceutical and Biomedical Analysis·Yukihiro KurodaKatsumi Matsuzaki
Jul 14, 2006·Electrophoresis·Junhua WangJieke Cheng
Aug 24, 2006·Electrophoresis·Yu-Fen HuangHuan-Tsung Chang
Dec 25, 2008·Electrophoresis·Kiyohito Shimura
Jan 20, 2004·Proteomics·Rulin ZhangGeorge Jackowski
Nov 3, 2020·Journal of Separation Science·Shannon L ThomasKatarína Maráková

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