Profiling of N-Linked Oligosaccharides of a Glycoprotein by UPLC-FLR-ESI-MS After Derivatization with Fluorescent Anthranilamide

Methods Mol Biol. 2021:2271:179-188. doi: 10.1007/978-1-0716-1241-5_13.

Abstract

N-glycans are described to have a large influence on the properties of therapeutic proteins, including safety and efficacy. For this reason, the extent and type of glycosylation is a characterization parameter for the analysis of antibodies and other therapeutic proteins. The method described here is a fast and high-throughput method for identification and semiquantification of N-glycans by HILIC-FLR-ESI-MS. Sample preparation has been optimized and simultaneous preparation of a large number of samples can be achieved within a day. The use of MS coupled to fluorescence detection is an additional tool for identifying the N-glycan type.

Keywords: HILIC; Mass spectrometry; Monoclonal antibodies; N-glycan.

MeSH terms

  • Antibodies, Monoclonal / analysis*
  • Chromatography, Liquid*
  • Fluorescent Dyes / chemistry*
  • Fluorometry
  • Glycoproteins / analysis*
  • Glycosylation
  • High-Throughput Screening Assays
  • Protein Processing, Post-Translational*
  • Research Design
  • Spectrometry, Mass, Electrospray Ionization*
  • Workflow
  • ortho-Aminobenzoates / chemistry*

Substances

  • Antibodies, Monoclonal
  • Fluorescent Dyes
  • Glycoproteins
  • ortho-Aminobenzoates
  • anthranilamide