Study_9S

Design, Implementation and Multisite Evaluation of a System Suitability Protocol for the Quantitative Assessment of Instrument Performance in Liquid Chromatography-Multiple Reaction Monitoring-MS (LC-MRM-MS)

  • Organism: Bos taurus
  • Instrument: TSQ Quantum,TSQ Vantage,4000 QTRAP,QTRAP 5500,6410 Triple Quadrupole LC/MS,6460 Triple Quadrupole LC/MS,Xevo TQ-XS
  • SpikeIn: No
  • Keywords: mass spectrometry, quantification, selected reaction monitoring, system suitability, skyline
Abstract
Multiple reaction monitoring (MRM) mass spectrometry coupled with stable isotope dilution (SID) and liquid chromatography (LC) is increasingly used in biological and clinical studies for precise and reproducible quantification of peptides and proteins in complex sample matrices. Robust LC-SID-MRM-MS-based assays that can be replicated across laboratories and ultimately in clinical laboratory settings require standardized protocols to demonstrate that the analysis platforms are performing adequately. We developed a system suitability protocol (SSP), which employs a predigested mixture of six proteins, to facilitate performance evaluation of LC-SID-MRM-MS instrument platforms, configured with nanoflow-LC systems interfaced to triple quadrupole mass spectrometers. The SSP was designed for use with low multiplex analyses as well as high multiplex approaches when software-driven scheduling of data acquisition is required. Performance was assessed by monitoring of a range of chromatographic and mass spectrometric metrics including peak width, chromatographic resolution, peak capacity, and the variability in peak area and analyte retention time (RT) stability. The SSP, which was evaluated in 11 laboratories on a total of 15 different instruments, enabled early diagnoses of LC and MS anomalies that indicated suboptimal LC-MRM-MS performance. The observed range in variation of each of the metrics scrutinized serves to define the criteria for optimized LC-SID-MRM-MS platforms for routine use, with pass/fail criteria for system suitability performance measures defined as peak area coefficient of variation <0.15, peak width coefficient of variation <0.15, standard deviation of RT <0.15 min (9 s), and the RT drift <0.5min (30 s). The deleterious effect of a marginally performing LC-SID-MRM-MS system on the limit of quantification (LOQ) in targeted quantitative assays illustrates the use and need for a SSP to establish robust and reliable system performance. Use of a SSP helps to ensure that analyte quantification measurements can be replicated with good precision within and across multiple laboratories and should facilitate more widespread use of MRM-MS technology by the basic biomedical and clinical laboratory research communities
Created on 1/22/18 10:28 PM
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Study9S_Site95_v1_2013-01-29_11-33-23.zip (3 MB)2018-01-227222311513
Study9S_Site98_v1_2013-01-29_11-39-10.zip (3 MB)2018-01-227222311513
Study9S_Site19_v1_2013-01-29_10-19-25.zip (3 MB)2018-01-227222311513
Study9S_Site20_v1_2013-01-29_10-25-32.zip (18 MB)2018-01-226222211013
Study9S_Site32_v1_2013-01-29_10-56-12.zip (5 MB)2018-01-227222311513
Study9S_Site52_v1_2013-01-29_10-06-21.zip (3 MB)2018-01-227222311513
Study9S_Site54_v1_2013-01-29_10-13-10.zip (4 MB)2018-01-227222311513
Study9S_Site56_v1_2013-01-29_11-00-30.zip (3 MB)2018-01-227222311513
Study9S_Site56A_v1_2013-01-29_11-04-41.zip (4 MB)2018-01-227222311513
Study9S_Site56B_v1_2013-01-29_11-08-46.zip (7 MB)2018-01-227222211013
Study9S_Site65_v1_2013-01-29_11-12-27.zip (17 MB)2018-01-226222211013
Study9S_Site73_v1_2013-01-29_11-14-57.zip (3 MB)2018-01-227222311513
Study9S_Site86_v1_2013-01-29_11-22-57.zip (4 MB)2018-01-227222311513
Study9S_Site86a_v1_2013-01-29_11-26-37.zip (7 MB)2018-01-227222311513
Study9S_Site90_v1_2013-01-29_11-30-59.zip (10 MB)2018-01-227222211013