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Citation: YAN Meng, ZHAO Yang, ZHANG Yangjun, YING Wantao, QIAN Xiaohong. Establishment of a microscale peptide prefractionation system based on ultra performance liquid chromatography and the eight-port rotor valve[J]. Chinese Journal of Chromatography, ;2019, 37(5): 477-483. doi: 10.3724/SP.J.1123.2018.11021 shu

Establishment of a microscale peptide prefractionation system based on ultra performance liquid chromatography and the eight-port rotor valve

1.
College of Pharmacy, Guangdong Pharmaceutical University, Guangzhou 510006, China;
2.
State Key Laboratory of Proteomics, Beijing Proteome Research Center, National Center for Protein Sciences(Beijing), Beijing Institute of Lifeomics, Beijing 102206, China

Received Date: 19 November 2018

Fund Project: National Key Program for Basic Research of China (Nos. 2017YFC0906600, 2016YFA0501300)

Liquid chromatography (LC) and mass spectrometry (MS)-based proteomics now allows very deep coverage of proteomes. Two-dimensional high performance liquid chromatography (2-D HPLC) is a useful tool for the proteome analysis of complex biosystem. However, it has drawback of a long running time, and it typically requires peptide amounts in the milligram range, and large volume of collected fractions. In this study, we introduce ultra-performance liquid chromatography (UPLC) and an eight-port rotor valve as a highly efficient and convenient method for a first-dimension separation and collection system. The combination of our UPLC-based fractionation using basic buffers with an online LC-MS/MS provided orthogonal peptide separation and demonstrated the powerful performance for protein identification. Upon applying the novel method to triplicate measurements of a human cell line, we observed excellent quantitative reproducibility between replicates (coefficient of determination R²>0.95) and more than 23.52% peptide identifications over the conventional StageTip approach. The fractionation method described here is flexible, straightforward, and robust, and it enables proteome analysis with minimal sample requirements.
- two-dimensional liquid chromatography (2-D LC),
- ultra performance liquid chromatography (UPLC),
- the eight-port rotor valve,
- proteomics,
- fractionation
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