Robust, reproducible and quantitative analysis of thousands of proteomes by micro-flow LC-MS/MS

Bian, Y., Zheng, R., Bayer, F. P., Wong, C., Chang, Y. C., Meng, C., Zolg, D. P., Reinecke, M., Zecha, J., Wiechmann, S., Heinzlmeir, S., Scherr, J., Hemmer, B., Baynham, M., Gingras, A. C., Boychenko, O., and Kuster, B. (2020). Nat Commun 11, 157. doi: 10.1038/s41467-019-13973-x


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.