Comparison of depletion strategies for the enrichment of low-abundance proteins in urine

Filip, S., Vougas, K., Zoidakis, J., Latosinska, A., Mullen, W. , Spasovski, G., Mischak, H., Vlahou, A. and Jankowski, J. (2015) Comparison of depletion strategies for the enrichment of low-abundance proteins in urine. PLoS ONE, 10(7), e0133773. (doi: 10.1371/journal.pone.0133773) (PMID:26208298) (PMCID:PMC4514849)

[img]
Preview
Text
109198.pdf - Published Version
Available under License Creative Commons Attribution.

2MB

Abstract

Proteome analysis of complex biological samples for biomarker identification remains challenging, among others due to the extended range of protein concentrations. High-abundance proteins like albumin or IgG of plasma and urine, may interfere with the detection of potential disease biomarkers. Currently, several options are available for the depletion of abundant proteins in plasma. However, the applicability of these methods in urine has not been thoroughly investigated. In this study, we compared different, commercially available immunodepletion and ion-exchange based approaches on urine samples from both healthy subjects and CKD patients, for their reproducibility and efficiency in protein depletion. A starting urine volume of 500 μL was used to simulate conditions of a multi-institutional biomarker discovery study. All depletion approaches showed satisfactory reproducibility (n=5) in protein identification as well as protein abundance. Comparison of the depletion efficiency between the unfractionated and fractionated samples and the different depletion strategies, showed efficient depletion in all cases, with the exception of the ion-exchange kit. The depletion efficiency was found slightly higher in normal than in CKD samples and normal samples yielded more protein identifications than CKD samples when using both initial as well as corresponding depleted fractions. Along these lines, decrease in the amount of albumin and other targets as applicable, following depletion, was observed. Nevertheless, these depletion strategies did not yield a higher number of identifications in neither the urine from normal nor CKD patients. Collectively, when analyzing urine in the context of CKD biomarker identification, no added value of depletion strategies can be observed and analysis of unfractionated starting urine appears to be preferable.

Item Type:Articles
Status:Published
Refereed:Yes
Glasgow Author(s) Enlighten ID:Mullen, Dr Bill and Mischak, Professor Harald
Authors: Filip, S., Vougas, K., Zoidakis, J., Latosinska, A., Mullen, W., Spasovski, G., Mischak, H., Vlahou, A., and Jankowski, J.
College/School:College of Medical Veterinary and Life Sciences > Institute of Cardiovascular and Medical Sciences
Journal Name:PLoS ONE
Publisher:Public Library of Science
ISSN:1932-6203
ISSN (Online):1932-6203
Copyright Holders:Copyright © 2015 The Authors
First Published:First published in PLoS One 10(7):e0133773
Publisher Policy:Reproduced under a Creative Commons License

University Staff: Request a correction | Enlighten Editors: Update this record