Abstract
Questions concerning longitudinal data quality and reproducibility of proteomic laboratories spurred the Protein Research Group of the Association of Biomolecular Resource Facilities (ABRF-PRG) to design a study to systematically assess the reproducibility of proteomic laboratories over an extended period of time. Developed as an open study, initially 64 participants were recruited from the broader mass spectrometry community to analyze provided aliquots of a six bovine protein tryptic digest mixture every month for a period of nine months. Data were uploaded to a central repository, and the operators answered an accompanying survey. Ultimately, 45 laboratories submitted a minimum of eight LC-MSMS raw data files collected in data-dependent acquisition (DDA) mode. No standard operating procedures were enforced; rather the participants were encouraged to analyze the samples according to usual practices in the laboratory. Unlike previous studies, this investigation was not designed to compare laboratories or instrument configuration, but rather to assess the temporal intralaboratory reproducibility. The outcome of the study was reassuring with 80% of the participating laboratories performing analyses at a medium to high level of reproducibility and quality over the 9-month period. For the groups that had one or more outlying experiments, the major contributing factor that correlated to the survey data was the performance of preventative maintenance prior to the LC-MSMS analyses. Thus, the Protein Research Group of the Association of Biomolecular Resource Facilities recommends that laboratories closely scrutinize the quality control data following such events. Additionally, improved quality control recording is imperative. This longitudinal study provides evidence that mass spectrometry-based proteomics is reproducible. When quality control measures are strictly adhered to, such reproducibility is comparable among many disparate groups. Data from the study are available via ProteomeXchange under the accession code PXD002114.
Highlights
Assessing Longitudinal Variability in LC-MSMS nity continues to exponentially grow and expand
Perhaps the most systematic and detailed of these investigations are from the Human Proteome Organization (HuPO) test sample working group (3); the National Cancer Institute Clinical Proteomic Tumor Analysis Consortium (NCI CPTAC) (4); and the ProteoRed Consortium (5, 6)
By applying the performance metrics developed by Rudnick et al (7), several key points emerged: (1) as expected, intralaboratory variation was less than interlaboratory variation; and (2) overall, the interlaboratory variation in peptide identifications and some of the other performance metrics were comparable between instruments, The abbreviations used are: LC-MSMS, liquid chromatography tandem mass spectrometry; DDA, data-dependent acquisition; FDR, false-discovery rate; HPLC, high-performance liquid chromatography; IQR, interquartile range.; LCMS, liquid chromatography mass spectrometry; MSMS, tandem mass spectrometry; PCA, principal component analysis; PM, preventative maintenance; Peptide-spectrum matches (PSMs), peptidespectrum match; QC, quality control; SOP, standard operating procedure
Summary
The 2012/2013 ABRF Proteomic Research Group Study: Assessing Longitudinal Intralaboratory Variability in Routine Peptide Liquid Chromatography Tandem Mass Spectrometry Analyses*□S. No standard operating procedures were enforced; rather the participants were encouraged to analyze the samples according to usual practices in the laboratory Unlike previous studies, this investigation was not designed to compare laboratories or instrument configuration, but rather to assess the temporal intralaboratory reproducibility. The ProteoRed Consortium initiated the ProteoRed Multicenter Experiment for Quality Control (PMEQC) (5, 6) This longitudinal QC multicenter study involved 12 institutes, and was designed to assess: (1) intralaboratory repeatability of LC-MSMS proteomic data; (2) interlaboratory reproducibility; and (3) reproducibility across multiple instrument platforms. The HuPO plasma proteome project (HuPO PPP) distributed 20 human samples (five serum plus 3 ϫ 5 plasma samples treated with three different anticoagulants) to 35 laboratories spanning 13 countries (8) The purpose of this large-scale study was not to assess reproducibility per se, but rather to generate the largest and most comprehensive data set on the protein composition of human plasma/serum. This community has endeavored to provide tools to assess proteomic data quality, and this study provides additional insight into the application of such tools and the quality of data within respective laboratories
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
