Scientific Publications

Design, Implementation, and Multi-Site Evaluation of a System Suitability Protocol for the Quantitative Assessment of Instrument Performance in LC-MRM-MS.

Publication TypeJournal Article
AuthorsAbbatiello, SE, Mani D. R., Schilling B., Maclean B., Zimmerman LJ, Feng X., Cusack MP, Sedransk N., Hall SC, Addona T., Allen S., Dodder NG, Ghosh M., Held JM, Hedrick V., Inerowicz HD, Jackson A., Keshishian H., Kim JW, Lyssand JS, Riley CP, Rudnick P., Sadowski P., Shaddox K., Smith D., Tomazela D., Wahlander A., Waldemarson S., Whitwell CA, You J., Zhang S., Kinsinger CR, Mesri M., Rodriguez H., Borchers CH, Buck C., Fisher SJ, Gibson BW, Liebler D., Maccoss M., Neubert TA, Paulovich A., Regnier F., Skates SJ, Tempst P., Wang M., and Carr SA
AbstractMultiple reaction monitoring (MRM) mass spectrometry (MS) 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 eleven laboratories on a total of fifteen different instruments, enabled early diagnoses of LC and MS anomalies that indicated sub-optimal 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 (CV) <0.15, peak width CV <0.15, standard deviation of RT <0.15 min (9 sec) and the RT drift <0.5min (30 sec). The deleterious effect of a marginally performing LC-SID-MRM-MS system on the limit of quantification (LOQ) in targeted quantitative assays illustrates the utility and need for a SSP to establish robust and reliable system performance. Use of a SSP helps to insure that analyte quantification measurements can be replicated with good precision within and across multiple laboratories and should facilitate more widespread utilization of MRM-MS technology by the basic biomedical and clinical laboratory research communities.
Year of Publication2013
JournalMolecular & cellular proteomics : MCP
Date Published (YYYY/MM/DD)2013/05/20
ISSN Number1535-9476
DOI10.1074/mcp.M112.027078
PubMedhttp://www.ncbi.nlm.nih.gov/pubmed/23689285?dopt=Abstract