Chip-based reversed-phase liquid chromatography-mass spectrometry of permethylated N-linked glycans: A potential methodology for cancer-biomarker discovery

William R. Alley, Milan Madera, Yehia Mechref, Milos V. Novotny

Research output: Contribution to journalArticlepeer-review

135 Scopus citations

Abstract

The study of protein glycosylation in biological fluids and tissues has substantial medical importance, as changes in glycan structures have now been associated with a number of diseases. Quantification of glycomic-profile changes is becoming increasingly important in the search for disease biomarkers. Here, we report a highly reproducible combination of a glycomic sample preparation/solid-phase derivatization of glycoprotein-derived N-linked glycans with their subsequent microchip-based separation and mass-spectrometric (MS) measurements. Following our previously described reductive β-elimination for O-linked glycans with ammonia-borane complex to reduce N-linked structures, the N-linked alditol structures are effectively methylated in dimethylformamide medium to avoid artefacts in MS measurements. Reversed-phase microfluidic liquid chromatography (LC) of methylated N-linked oligosaccharide alditols resolved some closely related structures into regular retention increments, aiding in their structural assignments. Optimized LC gradients, together with nanospray MS, have been applied here in the quantitative measurements of N-linked glycans in blood serum, distinguishing breast cancer patients from control individuals.

Original languageEnglish
Pages (from-to)5095-5106
Number of pages12
JournalAnalytical Chemistry
Volume82
Issue number12
DOIs
StatePublished - Jun 15 2010

Fingerprint

Dive into the research topics of 'Chip-based reversed-phase liquid chromatography-mass spectrometry of permethylated N-linked glycans: A potential methodology for cancer-biomarker discovery'. Together they form a unique fingerprint.

Cite this