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High-pH Reversed-Phase Pre-Fractionation for In-Depth Shotgun Proteomics

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Neuroproteomics

Part of the book series: Neuromethods ((NM,volume 146))

Abstract

Two-dimensional LC-MS-based proteomics is a powerful strategy for high-throughput protein identification and quantification. The combination of high-pH reversed-phase fractionation with low-pH LC-MS analysis is the method of choice, because it provides the optimal separation efficiency and deep protein coverage. In this chapter, we illustrate the protocols of HPLC-based and the tip-based high-pH reversed-phase chromatographic fractionation of peptide mixtures, in which the HPLC-based reversed-phase fractionation provides higher resolving power while the tip-based high-pH reversed-phase fractionation is simple, parallelizable, and suitable for limited amount of biological samples.

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Author information

Author notes

  1. Ning Chen and Mingwei Liu contributed equally to this work and are co-first authors.

Authors and Affiliations

  1. State Key Laboratory of Proteomics, Beijing Proteome Research Center, National Center for Protein Sciences Beijing, Beijing Institute of Lifeomics, Beijing, China

    Ning Chen, Mingwei Liu, Jun Qin, Wei Sun & Fuchu He

Authors
  1. Ning Chen
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  2. Mingwei Liu
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  3. Jun Qin
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  4. Wei Sun
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  5. Fuchu He
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Corresponding authors

Correspondence to Wei Sun or Fuchu He .

Editor information

Editors and Affiliations

  1. Department of Molecular and Cellular Neurobiology, Center for Neurogenomics and Cognitive Research, Amsterdam Neuroscience, Vrije Universiteit Amsterdam, Amsterdam, The Netherlands

    Ka Wan Li

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Chen, N., Liu, M., Qin, J., Sun, W., He, F. (2019). High-pH Reversed-Phase Pre-Fractionation for In-Depth Shotgun Proteomics. In: Li, K. (eds) Neuroproteomics. Neuromethods, vol 146. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-9662-9_5

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  • DOI: https://doi.org/10.1007/978-1-4939-9662-9_5

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-4939-9661-2

  • Online ISBN: 978-1-4939-9662-9

  • eBook Packages: Springer Protocols

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