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Conformational Analysis of Recombinant Monoclonal Antibodies with Hydrogen/Deuterium Exchange Mass Spectrometry

Protocol
First Online:
Part of the Methods in Molecular Biology book series (MIMB, volume 988)

Abstract

Understanding the conformation of antibodies, especially those of therapeutic value, is of great interest. Many of the current analytical methods used to probe protein conformation face issues in the analysis of antibodies, either due to the nature of the antibody itself or due to the limitations of the method. One method that has recently been utilized for conformational analysis of antibodies is hydrogen/deuterium exchange mass spectrometry (H/DX MS). H/DX MS can be used to probe the conformation and dynamics of proteins in solution, requires small sample quantities, is compatible with many buffer systems, and provides peptide-level resolution. The application of H/DX MS to immunoglobulin gamma 1 (IgG1) recombinant monoclonal antibodies can provide information about IgG1 conformation, dynamics, and changes to conformation as a result of protein modification(s), changes in storage conditions, purification procedures, formulation, and many other parameters. In this article we provide a comprehensive H/DX MS protocol for the analysis of an antibody.

Key words

Immunoglobin Deuterium Conformation Protein structure Antibody Hydrogen exchange 
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Notes

Acknowledgments

We would like to thank Dr. Steven A. Berkowitz for his advice and encouragement, Dr. Rohin Mhatre, Dr. Helena Madden, and Dr. Geoff Gerhardt for their support of H/DX MS, and Dr. Thomas E. Wales, Dr. Keith Fadgen, Dr. Martha Stapels, and Dr. Michael Eggertson for their technical assistance. This work was supported in part by funding from the NIH (GM-086507) and a research collaboration with Waters Corporation. This is contribution 974 from the Barnett Institute.

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Copyright information

© Springer Science+Business Media New York 2013

Authors and Affiliations

  1. 1.Biogen Idec, Inc.CambridgeUSA
  2. 2.Department of Chemistry and Chemical Biology, The Barnett Institute of Chemical and Biological AnalysisNortheastern UniversityBostonUSA

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