Microscale depletion of high abundance proteins in human biofluids using IgY14 immunoaffinity resin [electronic resource] : Analysis of human plasma and cerebrospinal fluid

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Tác giả:

Ngôn ngữ: eng

Ký hiệu phân loại: 572.7 Enzymes

Thông tin xuất bản: Richland, Wash. : Oak Ridge, Tenn. : Pacific Northwest National Laboratory (U.S.) ; Distributed by the Office of Scientific and Technical Information, U.S. Dept. of Energy, 2014

Mô tả vật lý: Size: p. 7117-7125 : , digital, PDF file.

Bộ sưu tập: Metadata

ID: 261370

 Removal of highly abundant proteins in plasma is often carried out using immunoaffinity depletion to extend the dynamic range of measurements to lower abundance species. While commercial depletion columns are available for this purpose, they generally are not applicable to limited sample quantities (<
 20 �L) due to low yields stemming from losses caused by nonspecific binding to the column matrix. Additionally, the cost of the depletion media can be prohibitive for larger scale studies. Modern LC-MS instrumentation provides the sensitivity necessary to scale-down depletion methods with minimal sacrifice to proteome coverage, which makes smaller volume depletion columns desirable for maximizing sample recovery when samples are limited, as well as for reducing the expense of large scale studies. We characterized the performance of a 346 �L column volume micro-scale depletion system, using four different flow rates to determine the most effective depletion conditions for ~6 ?L injections of human plasma proteins and then evaluated depletion reproducibility at the optimum flow rate condition. Depletion of plasma using a commercial 10 mL depletion column served as the control. Results showed depletion efficiency of the micro-scale column increased as flow rate decreased, and that our micro-depletion was reproducible. We found, in an initial application, a 600 �L sample of human cerebral spinal fluid (CSF) pooled from multiple sclerosis patients was depleted and then analyzed using reversed phase liquid chromatography-mass spectrometry to demonstrate the utility of the system for this important biofluid where sample quantities are more commonly limited.
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