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An HS-MRM Assay for the Quantification of Host-cell Proteins in Protein Biopharmaceuticals by Liquid Chromatography Ion Mobility QTOF Mass Spectrometry
11:09

An HS-MRM Assay for the Quantification of Host-cell Proteins in Protein Biopharmaceuticals by Liquid Chromatography Ion Mobility QTOF Mass Spectrometry

Published on: April 17, 2018

Development of an analytical reversed-phase high-performance liquid chromatography-electrospray ionization mass

Thomas M Dillon1, Pavel V Bondarenko, Margaret Speed Ricci

  • 1Amgen Inc., Pharmaceutics Department, One Amgen Center Dr, Thousand Oaks, CA 91320, USA.

Journal of Chromatography. A
|November 17, 2004
PubMed

Analytical characterization of monoclonal antibodies has been hindered by the lack of appropriate chromatographic methods to be used in conjunction with high-resolution MS. Current methodologies for standard RP-HPLC are incompatible with antibodies due to irreproducibility, low recovery, short column lifetimes, and poor resolution of degradation products. An analytical RP-HPLC-MS method was developed for monitoring and characterizing intact IgG1antibodies. Key parameters required for improved chromatographic resolution included long alkyl chains of the stationary phase (Zorbax SB300 C8), column temperatures elevated to 65-70 degrees C and combination of trifluoroacetic acid and heptafluorobutyric acid ion-pairing agents. RP chromatographic separation of degradation species and C-terminal lysine variants along with the characterization of glycosylation profile by mass spectrometry demonstrates the capability of this method for whole antibody analysis.

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