Serum Glycoproteome Profiling by Mass Spectrometry

Serum samples from volunteer donors were first enriched for 14 abundant serum proteins and digested with trypsin. Glycopeptides were enriched from the peptide mixture using either size exclusion chromatography or mixed-mode anion exchange cartridge (MAX), and analyzed by mass spectrometry (MS) in data dependent acquisition mode an Orbitrap Eclipse mass spectrometer (Thermo Fisher Scientific) [11 (link), 16 (link), 17 (link)]. Data was searched in pGlyco3 [18 (link)]. Commercial bovine (Thermo Scientific) and rabbit (Sigma) serum albumin were digested followed by glycopeptide enrichment using MAX. Details of sample preparation and MS analysis are provided in Additional file 1: Supplemental Methods.

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Garapati K., Jain A., Madden B.J., Mun D.G., Sharma J., Budhraja R, & Pandey A. (2024). Defining albumin as a glycoprotein with multiple N-linked glycosylation sites. Journal of Translational Medicine, 22, 454.

Publication 2024

Orbitrap Eclipse mass spectrometer

Corresponding Organization :

Other organizations : Institute of Bioinformatics, Mayo Clinic, Manipal Academy of Higher Education

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Variable analysis

independent variables

Enrichment method for glycopeptides (size exclusion chromatography or mixed-mode anion exchange cartridge (MAX))

dependent variables

Glycopeptide composition (as analyzed by mass spectrometry)

control variables

Serum samples from volunteer donors
Enrichment of 14 abundant serum proteins
Trypsin digestion of peptides
Mass spectrometry analysis in data dependent acquisition mode on an Orbitrap Eclipse mass spectrometer
Data search in pGlyco3

controls

Positive control: Commercial bovine serum albumin and rabbit serum albumin were digested and glycopeptides were enriched using MAX.
Negative control: Not explicitly mentioned.

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