Metrological traceability in mass spectrometry-based targeted protein quantitation: a proof-of-principle study for serum apolipoproteins A-I and B100.

Autor: Smit NP; Department of Clinical Chemistry and Laboratory Medicine, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, The Netherlands. Electronic address: N.Smit@lumc.nl., Romijn FP; Department of Clinical Chemistry and Laboratory Medicine, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, The Netherlands., van den Broek I; Department of Clinical Chemistry and Laboratory Medicine, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, The Netherlands., Drijfhout JW; Department of Immunohematology and Bloodtransfusion, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, The Netherlands., Haex M; Agilent Technologies Netherlands B.V., Amstelveen, The Netherlands., van der Laarse A; Department of Clinical Chemistry and Laboratory Medicine, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, The Netherlands., van der Burgt YE; Center for Proteomics and Metabolomics, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, The Netherlands., Cobbaert CM; Department of Clinical Chemistry and Laboratory Medicine, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, The Netherlands. Electronic address: C.M.Cobbaert@lumc.nl.
Jazyk: angličtina
Zdroj: Journal of proteomics [J Proteomics] 2014 Sep 23; Vol. 109, pp. 143-61. Date of Electronic Publication: 2014 Jun 25.
DOI: 10.1016/j.jprot.2014.06.015
Abstrakt: In this study, we have followed up on previous liquid chromatography (LC) multiple reaction monitoring (MRM) mass spectrometry (MS) approaches for measurement of apolipoprotein (apo) A-I and apo B100 in serum aiming for implementation of a multiplexed assay in a clinical chemistry laboratory with full metrological traceability. Signature peptides were selected and detected by dynamic MRM, and stable isotope labeled (SIL)-peptides were used as internal standards. Five apo A-I and four apo B100 peptides were measured in serum digests with linearity (R(2)>0.992) in the physiologically relevant concentration ranges. Linearity with regard to protein concentration was ascertained at five concentration levels (R(2)>0.926 and R(2)>0.965, for the apo A-I and apo B100 peptides, respectively). Three native value-assigned sera were used as external calibrators for further method verification. Imprecision values on sample preparation and LC-MS/MS acquisition were below the established minimal specifications for apo A-I and apo B100 (5.0% and 5.3%, respectively). Correlation of LC-MS/MS results with immunoturbidimetric assay results, for normo- and hypertriglyceridemic samples, showed R(2)>0.944 for apo A-I and R(2)>0.964 for apo B100. This LC-MS/MS method has potential for clinical application in normo- and dyslipidemic patients.
Biological Significance: Measurement of apo A-I and apo B100 may offer an alternative to high and low density lipoprotein cholesterol (HDL-c and LDL-c) methods for cardiovascular disease risk assessment in dyslipidemic patients [1]. An LC-MS/MS method for apo A-I and apo B100 has the advantage of antibody independent and specific detection of protein signature peptides. The introduction of an LC-MS/MS method for apo A-I and apo B100 can serve as an example for many existing and newly developed (multiplex) biomarker methods in quantitative clinical chemistry proteomics (qCCP). Such LC-MS/MS methods should meet basic clinical chemistry principles with regard to test evaluation [2]. Criteria for imprecision should be pre-defined, e.g., based on biological variation. The use of commutable and traceable serum-based calibrators will improve inter-laboratory reproducibility of LC-MS/MS methods and may contribute to a more rapid transition of biomarker discovery to clinical utility with benefit for the patient treatment and improvement of general health care.
(Copyright © 2014 Elsevier B.V. All rights reserved.)
Databáze: MEDLINE