Characterization of immunoglobulins through analysis of N-glycopeptides by MALDI-TOF MS

Methods : a Companion to Methods in Enzymology
Emy KomatsuHélène Perreault

Abstract

The aim of this report is to emphasize the role, usefulness and power of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) in the analysis of glycoforms of antibodies (Abs) through their proteolytic glycopeptides. Abs are complex biomolecules in which glycans hold determinant properties and thus need to be thoroughly characterized following Ab production by recombinant methods or Ab collection from human/animal serum or tissue. In spite of the great robustness of MALDI-TOF MS in terms of tolerance to impurities, the analysis of Abs and Ab components using this technique requires extensive sample preparation involving all or some of chromatography, solid phase extraction, enzymatic modification, and chemical derivatization. This report focuses on a monoclonal Ab produced in cell culture, as well as on a polyclonal human immunoglobulin (Ig) G obtained commercially and a polyclonal porcine IgG obtained from serum. A method is first provided to separate Ab protein chain components (light chains, heavy chains) by gel electrophoresis, which is useful for instance for protein-A eluates of Igs either from cell culture or biological samples. This allows for in-gel proteolytic digestion of the pr...Continue Reading

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Citations

Jun 22, 2016·Methods : a Companion to Methods in Enzymology·Rainer Cramer
Nov 21, 2017·SLAS Discovery·Dale S Cornett, Michael D Scholle
Nov 14, 2017·Expert Review of Proteomics·Róisín O'FlahertyGordan Lauc
Apr 5, 2016·Expert Review of Proteomics·Irena Trbojević-AkmačićGordan Lauc
Oct 6, 2017·JCI Insight·Adrien BosseboeufSylvie Hermouet
Jan 5, 2021·Frontiers in Plant Science·Lennard L BohlenderEva L Decker
Feb 5, 2021·Rapid Communications in Mass Spectrometry : RCM·Taylor BattellinoHélène Perreault
Feb 13, 2021·European Journal of Immunology·Bernard VanhoveJean-Paul Soulillou
May 26, 2018·Journal of the American Society for Mass Spectrometry·Rini RoyHélène Perreault

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