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Title: Microscale depletion of high abundance proteins in human biofluids using IgY14 immunoaffinity resin: Analysis of human plasma and cerebrospinal fluid

Journal Article · · Analytical and Bioanalytical Chemistry
 [1];  [2];  [2];  [2];  [2];  [2];  [2];  [2];  [2];  [2];  [2];  [2]
  1. Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Korea Research Inst. of Standards and Science (Korea)
  2. Pacific Northwest National Lab. (PNNL), Richland, WA (United States)
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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 µL) 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 µL column volume micro-scale depletion system, using four different flow rates to determine the most effective depletion conditions for ~6 μL 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 µL 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.

Research Organization:
Pacific Northwest National Laboratory (PNNL), Richland, WA (United States). Environmental Molecular Sciences Laboratory (EMSL)
Sponsoring Organization:
USDOE
Grant/Contract Number:
AC05-76RL01830; AC05-76RLO01830; 8 P41 GM103493-10; 5 P41 RR018522-10
OSTI ID:
1168898
Report Number(s):
PNNL-SA-92758; 47418; 40072; 48135; 400412000
Journal Information:
Analytical and Bioanalytical Chemistry, Vol. 406, Issue 28; ISSN 1618-2642
Publisher:
SpringerCopyright Statement
Country of Publication:
United States
Language:
English
Citation Metrics:
Cited by: 10 works
Citation information provided by
Web of Science

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Cited By (2)

Label-free quantitative proteomic analysis reveals potential biomarkers for early healing in cutaneous leishmaniasis journal January 2019
High-abundant protein depletion strategies applied on dog cerebrospinal fluid and evaluated by high-resolution mass spectrometry journal September 2015

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Related Subjects

60 APPLIED LIFE SCIENCES
59 BASIC BIOLOGICAL SCIENCES
microscale depletion
IgY-14 immunoaffinity resin
Human plasma
cerebrospinal fluid
MS
Environmental Molecular Sciences Laboratory