The utility of hydrogen/deuterium exchange mass spectrometry in biopharmaceutical comparability studies

J Pharm Sci. 2011 Jun;100(6):2071-86. doi: 10.1002/jps.22432. Epub 2010 Dec 29.


The function, efficacy, and safety of protein biopharmaceuticals are tied to their three-dimensional structure. The analysis and verification of this higher-order structure are critical in demonstrating manufacturing consistency and in establishing the absence of structural changes in response to changes in production. It is, therefore, essential to have reliable, high-resolution and high sensitivity biophysical tools capable of interrogating protein structure and conformation. Here, we demonstrate the use of hydrogen/deuterium exchange mass spectrometry (H/DX-MS) in biopharmaceutical comparability studies. H/DX-MS measurements can be conducted with good precision, consume only picomoles of protein, interrogate nearly the entire molecule with peptide level resolution, and can be completed in a few days. Structural comparability or lack of comparability was monitored for different preparations of interferon-β-1a. We present specific graphical formats for the display of H/DX-MS data that aid in rapidly making both the qualitative (visual) and quantitative assessment of comparability. H/DX-MS is capable of making significant contributions in biopharmaceutical characterization by providing more informative and confidant comparability assessments of protein higher-order structures than are currently available within the biopharmaceutical industry.

Publication types

  • Comparative Study
  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Biopharmaceutics / methods*
  • Biopharmaceutics / standards
  • Deuterium Exchange Measurement*
  • Interferons / chemistry
  • Mass Spectrometry*
  • Models, Molecular
  • Protein Conformation
  • Proteins / chemistry*
  • Technology, Pharmaceutical / methods*
  • Technology, Pharmaceutical / standards


  • Proteins
  • Interferons