High-throughput lectin magnetic bead array-coupled tandem mass spectrometry for glycoprotein biomarker discovery

Eun ju Choi, Dorothy Loo, James W. Dennis, Caroline A. O'Leary, Michelle M. Hill

Research output: Contribution to journalArticlepeer-review

30 Scopus citations

Abstract

Alterations in protein glycosylation occur during development and progression of many diseases, hence glycomics and glycoproteomics have emerged as important tools in glycobiomarker discovery. High-throughput glycan profiling can now be achieved with the recent developments in MS-based techniques. To enable identification and rapid monitoring of glycosylation changes in serum proteins, we developed a semi-automated high-throughput glycoprotein biomarker discovery platform termed lectin magnetic bead array-coupled tandem mass spectrometry (LeMBA-MS) which includes (i) effective single-step serum glycoprotein isolation using a panel of 20 individual lectin-coated magnetic beads in microplate format, (ii) on-bead trypsin digestion, and (iii) nanoLC-MS/MS with lectin exclusion list. With use of appropriate sequence databases, LeMBA-MS can detect glycosylation changes regardless of the species. By spiking known amounts of titrated ovalbumin to a serum sample, we report nanomolar sensitivity, and linearity of response of LeMBA-MS using concanavalin A-coupled beads. Neuraminidase treatment led to reduction of binding to sialic acid-binding lectins. Interestingly, we found that desialylation caused increased binding of haptoglobin and hemopexin to mannose-specific lectins, pointing to the importance of identifying a signature of lectin-binding. High-throughput LeMBA-MS to generate glycosylation signatures will facilitate glycobiomarker discovery. LeMBA can be coupled to down-stream detection platforms for validation, making it a truly versatile platform.

Original languageEnglish (US)
Pages (from-to)3564-3575
Number of pages12
JournalElectrophoresis
Volume32
Issue number24
DOIs
StatePublished - Dec 1 2011
Externally publishedYes

Keywords

  • Biomarker discovery
  • Glycoproteome fractionation
  • Glycosylation
  • Magnetic beads
  • Multilectin affinity

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry
  • Clinical Biochemistry

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