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Talanta. 2017 Mar 1;164:77-84. doi: 10.1016/j.talanta.2016.11.023. Epub 2016 Nov 13.

Multiplex and accurate quantification of acute kidney injury biomarker candidates in urine using Protein Standard Absolute Quantification (PSAQ) and targeted proteomics.

Author information

1
Université Grenoble-Alpes, F-38000 Grenoble, France; CEA, BIG, Biologie à Grande Echelle, F-38054 Grenoble, France; INSERM, U1038, F-38054 Grenoble, France.
2
AP-HP, Hôpital Tenon, Department of Nephrology and Dialysis, F-75020 Paris, France.
3
INSERM, UMR_S 1155, F-75005 Paris, France.
4
AP-HP, Hôpital Henri Mondor, Department of Nephrology, F-94010 Créteil, France.
5
AP-HP, Hôpital Henri Mondor, Plateforme de Ressources Biologiques, F-94010 Créteil, France.
6
AP-HP, Hôpital Tenon, Department of Nephrology and Dialysis, F-75020 Paris, France; INSERM, UMR_S 1155, F-75005 Paris, France; Sorbonne Universités, UPMC Univ Paris 06, UMR_S 1155, F-75005 Paris, France.
7
Université Grenoble-Alpes, F-38000 Grenoble, France; CEA, BIG, Biologie à Grande Echelle, F-38054 Grenoble, France; INSERM, U1038, F-38054 Grenoble, France. Electronic address: virginie.brun@cea.fr.

Abstract

There is a need for multiplex, specific and quantitative methods to speed-up the development of acute kidney injury biomarkers and allow a more specific diagnosis. Targeted proteomic analysis combined with stable isotope dilution has recently emerged as a powerful option for the parallelized evaluation of candidate biomarkers. This article presents the development of a targeted proteomic assay to quantify 4 acute kidney injury biomarker candidates in urine samples. The proteins included in the assessed panel consisted of myo-inositol oxygenase (MIOX), phosphoenolpyruvate carboxykinase 1 (PCK1), neutrophil gelatinase-associated lipocalin (NGAL) and liver fatty acid-binding protein (L-FABP). The proteomic assay combined an antibody-free sample preparation and a liquid chromatography-selected reaction monitoring (LC-SRM) analysis pipeline. For accurate quantification of the selected candidates, we used PSAQ (Protein Standard Absolute Quantification) standards which are isotopically labeled versions of the target proteins. When added directly to the biological samples, these standards improve detection specificity and quantification accuracy. The multiplexed assay developed for the 4 biomarker candidates showed excellent analytical performance, in line with the recommendations of health authorities. Tests on urine from two small patient cohorts and a group of healthy donors confirmed the relevance of NGAL and L-FABP as biomarkers for AKI diagnosis. The assay is readily adaptable to other biomarker candidates and should be very useful for the simultaneous and accurate quantification of multiple biomarkers.

KEYWORDS:

Biomarker; Kidney; Protein standard absolute quantification; Proteomics; Quantification; Selected reaction monitoring

PMID:
28107998
DOI:
10.1016/j.talanta.2016.11.023
[Indexed for MEDLINE]

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