Sample preparation scale-up for deep nglycomic analysis of human serum by capillary electrophoresis and CE-ESI-MS

Marton Szigeti, Andras Guttman

Research output: Contribution to journalArticle

1 Citation (Scopus)

Abstract

We introduce an efficient sample preparation workflow to facilitate deep N-glycomics analysis of the human serum by capillary electrophoresis with laser induced fluorescence (CE-LIF) detection and to accommodate the higher sample concentration requirement of electrospray ionization mass spectrometry connected to capillary electrophoresis (CE-ESI-MS). A novel, temperature gradient denaturing protocol was applied on amine functionalized magnetic bead partitioned glycoproteins to circumvent the otherwise prevalent precipitation issue. During this process, the free sugar content of the serum was significantly decreased as well, accommodating enhanced PNGase F mediated release of the N-linked carbohydrates. The liberated oligosaccharides were tagged with aminopyrene- trisulfonate, utilizing a modified evaporative labeling protocol. Processing the samples with this new workflow enabled deep CE-LIF analysis of the human serum N-glycome and provided the appropriate amount of material for CE-ESI-MS analysis in negative ionization mode.

Original languageEnglish
Pages (from-to)2524-2531
Number of pages8
JournalMolecular and Cellular Proteomics
Volume18
Issue number12
DOIs
Publication statusPublished - 2019

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry
  • Molecular Biology

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