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.
CITATION STYLE
Chen, N., Liu, M., Qin, J., Sun, W., & He, F. (2019). High-pH reversed-phase pre-fractionation for in-depth shotgun proteomics. In Neuromethods (Vol. 146, pp. 55–65). Humana Press Inc. https://doi.org/10.1007/978-1-4939-9662-9_5
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