Robust, reproducible and quantitative analysis of thousands of proteomes by micro-flow LC-MS/MS.
Nat Commun. 2020 Jan 9;11(1):157. doi: 10.1038/s41467-019-13973-x.
|Authors/Editors:||Bian Y, Zheng R, Bayer FP, Wong C, Chang YC, Meng C, Zolg DP, Reinecke M, Zecha J, Wiechmann S, Heinzlmeir S, Scherr J, Hemmer B, Baynham M, Gingras AC, Boychenko O, Kuster B.|
Nano-flow liquid chromatography tandem mass spectrometry (nano-flow LC–MS/MS) is the mainstay in proteome research because of its excellent sensitivity but often comes at the expense of robustness. Here we show that micro-flow LC–MS/MS using a 1 × 150 mm column shows excellent reproducibility of chromatographic retention time (<0.3% coefficient of variation, CV) and protein quantification (<7.5% CV) using data from >2000 samples of human cell lines, tissues and body fluids. Deep proteome analysis identifies >9000 proteins and >120,000 peptides in 16 h and sample multiplexing using tandem mass tags increases throughput to 11 proteomes in 16 h. The system identifies >30,000 phosphopeptides in 12 h and protein-protein or protein-drug interaction experiments can be analyzed in 20 min per sample. We show that the same column can be used to analyze >7500 samples without apparent loss of performance. This study demonstrates that micro-flow LC–MS/MS is suitable for a broad range of proteomic applications.