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Gstöttner, C., Reusch, D., Haberger, M., Irina, D., Van Veelen, P., Kilgour, D.P.A. 
ORCID: 0000-0002-3860-7532, Tsybin, Y.O., Van der Burgt, Y., Wuhrer, M. and Nicolardi, S.,
2020.
Monitoring glycation levels of a bispecific monoclonal antibody at subunit level by ultrahigh resolution MALDI FT-ICR mass spectrometry.
mAbs, 12 (1): e1682403.
ISSN 1942-0862
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1202258_Kilgour_a417.pdf - Published version Download (4MB) | Preview |
Abstract
Bispecific monoclonal antibodies (BsAbs) are engineered proteins with multiple functionalities and properties. The "bi-specificity" of these complex biopharmaceuticals is a key characteristic for the development of novel and more effective therapeutic strategies. The high structural complexity of BsAbs poses a challenge to the analytical methods needed for their characterization. Modifications of the BsAb structure, resulting from enzymatic and non-enzymatic processes, further complicate the analysis. An important example of the latter type of modification is glycation, which can occur in the manufacturing process, during storage in formulation or in vivo after application of the drug. Glycation affects the structure, function and stability of monoclonal antibodies, and consequently, detailed analysis of glycation levels is required. Mass spectrometry (MS) plays a key role in the structural characterization of monoclonal antibodies and top-down, middle-up and middle-down MS approaches are increasingly used for the analysis of modifications. Here, we apply a novel middle-up strategy, based on IdeS digestion and matrix-assisted laser desorption ionization (MALDI) Fourier transform ion cyclotron resonance (FT-ICR) MS, to analyze all six different BsAb subunits in a single high-resolution mass spectrum, namely two light chains, two half fragment crystallizable regions and two Fd’ regions, thus avoiding upfront chromatography. This method was used to monitor glycation changes during a 168h forced-glycation experiment. In addition, hot spot glycation sites were localized using top-down and middle-down MALDI-in-source decay FT-ICR MS, which provided complementary information compared to standard bottom-up MS.
| Item Type: | Journal article | ||||||
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| Publication Title: | mAbs | ||||||
| Creators: | Gstöttner, C., Reusch, D., Haberger, M., Irina, D., Van Veelen, P., Kilgour, D.P.A., Tsybin, Y.O., Van der Burgt, Y., Wuhrer, M. and Nicolardi, S. | ||||||
| Publisher: | Taylor and Francis | ||||||
| Date: | 2020 | ||||||
| Volume: | 12 | ||||||
| Number: | 1 | ||||||
| ISSN: | 1942-0862 | ||||||
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| Rights: | © 2019 The Author(s). Published with license by Taylor & Francis Group, LLC. This is an Open Access article distributed under the terms of the Creative Commons Attribution-NonCommercial License (http://creativecommons.org/licenses/by-nc/4.0/), which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited. | ||||||
| Divisions: | Schools > School of Science and Technology | ||||||
| Record created by: | Linda Sullivan | ||||||
| Date Added: | 26 Nov 2019 10:49 | ||||||
| Last Modified: | 27 Feb 2020 16:27 | ||||||
| URI: | https://irep.ntu.ac.uk/id/eprint/38508 |
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