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Protocols for the analytical characterization of therapeutic monoclonal antibodies. II - Enzymatic and chemical sample preparation.
Journal article

Protocols for the analytical characterization of therapeutic monoclonal antibodies. II - Enzymatic and chemical sample preparation.

  • Bobaly B School of Pharmaceutical Sciences, University of Geneva, University of Lausanne, Centre Médical Universitaire (CMU), Rue Michel-Servet 1, 1206, Geneva, Switzerland.
  • D'Atri V School of Pharmaceutical Sciences, University of Geneva, University of Lausanne, Centre Médical Universitaire (CMU), Rue Michel-Servet 1, 1206, Geneva, Switzerland.
  • Goyon A School of Pharmaceutical Sciences, University of Geneva, University of Lausanne, Centre Médical Universitaire (CMU), Rue Michel-Servet 1, 1206, Geneva, Switzerland.
  • Colas O Institut de Recherche Pierre Fabre, Centre d'Immunologie, 5 Avenue Napoléon III, BP 60497, 74160, Saint-Julien-en-Genevois, France.
  • Beck A Institut de Recherche Pierre Fabre, Centre d'Immunologie, 5 Avenue Napoléon III, BP 60497, 74160, Saint-Julien-en-Genevois, France.
  • Fekete S School of Pharmaceutical Sciences, University of Geneva, University of Lausanne, Centre Médical Universitaire (CMU), Rue Michel-Servet 1, 1206, Geneva, Switzerland.
  • Guillarme D School of Pharmaceutical Sciences, University of Geneva, University of Lausanne, Centre Médical Universitaire (CMU), Rue Michel-Servet 1, 1206, Geneva, Switzerland. Electronic address: davy.guillarme@unige.ch.
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  • 2017-07-01
Published in:
  • Journal of chromatography. B, Analytical technologies in the biomedical and life sciences. - 2017
Enzymatic digestion
Protocol
Reduction
Sample preparation
Therapeutic monoclonal antibodies
Antibodies, Monoclonal
Chromatography, High Pressure Liquid
Glycosylation
Mass Spectrometry
Papain
Peptide Fragments
Polysaccharides
Trypsin
English The analytical characterization of therapeutic monoclonal antibodies and related proteins usually incorporates various sample preparation methodologies. Indeed, quantitative and qualitative information can be enhanced by simplifying the sample, thanks to the removal of sources of heterogeneity (e.g. N-glycans) and/or by decreasing the molecular size of the tested protein by enzymatic or chemical fragmentation. These approaches make the sample more suitable for chromatographic and mass spectrometric analysis. Structural elucidation and quality control (QC) analysis of biopharmaceutics are usually performed at intact, subunit and peptide levels. In this paper, general sample preparation approaches used to attain peptide, subunit and glycan level analysis are overviewed. Protocols are described to perform tryptic proteolysis, IdeS and papain digestion, reduction as well as deglycosylation by PNGase F and EndoS2 enzymes. Both historical and modern sample preparation methods were compared and evaluated using rituximab and trastuzumab, two reference therapeutic mAb products approved by Food and Drug Administration (FDA) and European Medicines Agency (EMA). The described protocols may help analysts to develop sample preparation methods in the field of therapeutic protein analysis.
Language
  • English
Open access status
closed
Identifiers
Persistent URL
https://sonar.ch/global/documents/43663
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