A HUPO test sample study reveals common problems in mass spectrometry-based proteomics

Alexander W. Bell, Eric W. Deutsch, Gavin Meredith, Juan Antonio Vizca�no, Peter Sheffield, Phillip Gray, Mahbod Hajivandi, Marshall Pope, Paul Predki, Majlinda Kullolli, Maria Hincapie, William Stephen Hancock, Wei Jia, Catherine E. Au, Lina Song, Lei Li, Junying Wei, Bing Yang, Jinglan Wang, Wantao YingYangjun Zhang, Yun Cai, Xiaohong Qian, Fuchu He, Robert E. Kearney, Helmut Meyer, Christian Stephan, Martin Eisenacher, Katrin Marcus, Elmar Langenfeld, Caroline May, Steven A. Carr, Rushdy Ahmad, Wenhong Zhu, Jeffrey W. Smith, Ron Beavis, Samir M. Hanash, Jason J. Struthers, Hong Wang, Qing Zhang, Yanming An, Radoslav Goldman, Elisabet Carlsohn, Sjoerd van der Post, Kenneth E. Hung, David A. Sarracino, Salvatore Sechi, Kenneth Parker, Bryan Krastins, Raju Kucherlapati, Sylvie Bourassa, Guy G. Poirier, Eugene A. Kapp, Heather Patsiouras, Robert L. Moritz, Richard J. Simpson, Benoit Houle, Thomas Nilsson, Sylvie LaBoissiere, Pavel Metalnikov, Vivian Nguyen, Tony Pawson, Catherine C. L. Wong, Daniel Cociorva, J.R. Yates, Ana Lopez-Campistrous, Paul Semchuk, Yueju Wang, John J. M. Bergeron, Peipei Ping, Giuliano Elia, Michael J. Dunn, Kieran Wynne, Angela K. Walker, John R. Strahler, Philip Andrews, Brian L. Hood, William L. Bigbee, Thomas P. Conrads, Thomas A. Beardslee, Derek Smith, Christoph H. Borchers, Gilles A. Lajoie, Sean C. Bendall, Kaye D. Speicher, David W. Speicher, Masanori Fujimoto, Kazuyuki Nakamura, Young-Ki Paik, Sang Yun Cho, Thomas Chappell, Min-Seok Kwon, Hyoung-Joo Lee, Seul-Ki Jeong, An Sung Chung, Christine A. Miller, Rudolf Grimm, Katy Williams, Craig Dorschel, Jayson A. Falkner, Lennart Martens

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289 Citations (Scopus)


We performed a test sample study to try to identify errors leading to irreproducibility, including incompleteness of peptide sampling, in liquid chromatography-mass spectrometry-based proteomics. We distributed an equimolar test sample, comprising 20 highly purified recombinant human proteins, to 27 laboratories. Each protein contained one or more unique tryptic peptides of 1,250 Da to test for ion selection and sampling in the mass spectrometer. Of the 27 labs, members of only 7 labs initially reported all 20 proteins correctly, and members of only 1 lab reported all tryptic peptides of 1,250 Da. Centralized analysis of the raw data, however, revealed that all 20 proteins and most of the 1,250 Da peptides had been detected in all 27 labs. Our centralized analysis determined missed identifications (false negatives), environmental contamination, database matching and curation of protein identifications as sources of problems. Improved search engines and databases are needed for mass spectrometry-based proteomics.
Original languageEnglish
Pages (from-to)423-430
Number of pages8
JournalNature Methods
Issue number6
Publication statusPublished - 2009
Externally publishedYes


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