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Jooß, K. ; Hühner, J.* ; Kiessig, S.* ; Moritz, B.* ; Neusüß, C.*

Two-dimensional capillary zone electrophoresis–mass spectrometry for the characterization of intact monoclonal antibody charge variants, including deamidation products.

Anal. Bioanal. Chem. 409, 6057–6067 (2017)

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Open Access Green as soon as Postprint is submitted to ZB.

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Capillary zone electrophoresis (CZE) is a powerful tool that is progressively being applied for the separation of monoclonal antibody (mAb) charge variants. Mass spectrometry (MS) is the desired detection method concerning identification of mAb variants. In biopharmaceutical applications, there exist optimized and validated electrolyte systems for mAb variant quantification. However, these electrolytes interfere greatly with the electrospray ionization (ESI) process. Here, a heart-cut CZE–CZE–MS setup with an implemented mechanical four-port valve interface was developed that used a generic ε-aminocaproic acid based background electrolyte in the first dimension and acetic acid in the second dimension. Interference-free, highly precise mass data (deviation less than 1 Da) of charge variants of trastuzumab, acting as model mAb system, were achieved. The mass accuracy obtained (low parts per million range) is discussed regarding both measured and calculated masses. Deamidation was detected for the intact model antibody, and related mass differences were significantly confirmed on the deglycosylated level. The CZE–CZE–MS setup is expected to be applicable to a variety of antibodies and electrolyte systems. Thus, it has the potential to become a compelling tool for MS characterization of antibody variants separated in ESI-interfering electrolytes.

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Publication type Article: Journal article

Document type Scientific Article

Corresponding Author

Keywords Antibody Analysis ; Capillary Zone Electrophoresis–mass Spectrometry ; Electrospray Ionization ; Mechanical Valve ; Pharmaceutical Application ; Two-dimensional Interface ; ε-aminocaproic Acid; Biosimilar Rituximab; Mechanical Valve; Cze-ms; Identification; Chromatography; Separation; Heterogeneity; Trastuzumab; Recombinant; Proteins

ISSN (print) / ISBN 1618-2642

e-ISSN 1618-2650

Journal Analytical and Bioanalytical Chemistry

Quellenangaben Volume: 409, Issue: 26, Pages: 6057–6067

Publisher Springer

Publishing Place Heidelberg

Non-patent literature Publications

Reviewing status Peer reviewed

Institute(s) Research Unit Analytical BioGeoChemistry (BGC)