HPLC enrichment/isolation of proteins for post-translational modification studies from complex mixtures

J Pharm Biomed Anal. 2014 Sep:98:393-400. doi: 10.1016/j.jpba.2014.06.025. Epub 2014 Jun 21.

Abstract

The paper describes a macroporous RP-HPLC method for separation and isolation/enrichment of proteins from complex mixtures. The method is robust and efficient; using 2.1 or 4.6mm diameter columns provides sufficient material for subsequent proteomic analysis. The main advantage of the method is that most protein variants are isolated in the same fraction, as separation is not based on differences in isoelectric point. This is highly advantageous for studying complex mixtures and post-translational modifications. Examples related to glycosylation analysis are discussed in detail.

Keywords: Glycosylation; HPLC; Mass spectrometry; Proteins; Sample preparation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Chromatography, High Pressure Liquid / methods
  • Complex Mixtures / chemistry*
  • Glycosylation
  • Humans
  • Plasma / chemistry
  • Protein Processing, Post-Translational / genetics*
  • Proteins / chemistry*
  • Proteomics / methods

Substances

  • Complex Mixtures
  • Proteins