Glycosylation profiling of immunoglobulin G (IgG) subclasses from human serum

Proteomics. 2007 Nov;7(22):4070-81. doi: 10.1002/pmic.200700289.


All four subclasses of human serum IgG contain a single N-glycosylation site in the constant region of their heavy chain, which is occupied by biantennary, largely core-fucosylated and partially truncated oligosaccharides, that may carry a bisecting N-acetylglucosamine and sialic acid residues. IgG glycosylation has been shown to be altered under various physiological and pathological circumstances. IgG N-glycan profiles vary with age, and galactosylation for example is enhanced during pregnancy. Several diseases including rheumatoid arthritis are associated with a reduction in galactosylation of the IgG N-glycans. Here, we describe a robust method for the isolation of IgG subclasses using protein A (binds IgG1, IgG2, and IgG4) and protein G (binds additionally IgG3) at the 96-well plate level, which is suitable for automation. Isolated IgGs were digested with trypsin, and obtained glycopeptides were analyzed by nano-LC-MS. Glycopeptides were characterized by CID as well as electron transfer dissociation (ETD). The method provided glycosylation profiles for IgG1, IgG2, IgG3, and IgG4 and revealed distinct differences in N-glycosylation between the four IgG subclasses. The changes in galactosylation associated with rheumatoid arthritis could readily be monitored. This method is suitable for the subclass-specific analysis of IgG glycosylation from clinical samples.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Chromatography, Liquid / methods
  • Glycopeptides / analysis
  • Glycosylation
  • Humans
  • Immunoglobulin G* / blood
  • Immunoglobulin G* / classification
  • Immunoglobulin G* / immunology
  • Immunoglobulin Isotypes / blood
  • Immunoglobulin Isotypes / immunology
  • Immunoglobulin Isotypes / isolation & purification*
  • Proteomics / methods
  • Sensitivity and Specificity
  • Spectrometry, Mass, Electrospray Ionization / methods
  • Trypsin / chemistry


  • Glycopeptides
  • Immunoglobulin G
  • Immunoglobulin Isotypes
  • Trypsin