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Title: Specific enzymatic digestion and fragment isolation for the critical quality attribute assessment of IgG1
Authors : Rappo, Martin Alexander
Advisors / Reviewers : Rohrer, Jack
Rösli, Christoph
Extent : 57
Publisher / Ed. Institution : ZHAW Zürcher Hochschule für Angewandte Wissenschaften
Issue Date: 2017
License (according to publishing contract) : Licence according to publishing contract
Language : English
Subjects : Critical Quality Attributes; CQA; Monoclonal Antibody; Mab; Fab; Fc; F(ab’)2; Large hinge lgG fragment; LHF; IdeS; Lysine Gingipain; Kgp; EndoS2; IgdE
Subject (DDC) : 615: Pharmacology and therapeutics
660.6: Biotechnology
Abstract: Recent ICH quality guideline updates introduced the enhanced drug development which is based on Quality by Design approach. For this a Quality Risk Management has to be conducted based on scientific knowledge. Information gaps are closed through analysis of molecules with domain-specific modifications. The aim of this thesis was to evaluate methods to prepare a model IgG1 for such studies. The antibody fragments F(ab’)2, LHF, Fab and Fc and the Fc-glycosylation cleaved IgG with residual N-acetylglucosamine were selected. A representative IgG1 was digested with the recently discovered enzymes IdeS, Kgp, IgdE and EndoS2 and the digestion products were purified with common biotechnological down-stream methods. The F(ab’)2 was successfully isolated with a yield of 55% and a purity of 96%. After an initial digestion parameter optimization the LHF, Fab and Fc could be generated and an isolation method was established. A following scale-up was unsuccessful, due to reproducibility issues. A promising alternative was identified. The enzymatic glycan cleavage and product isolation was successful with a yield of 80% and a purity of 99%.
Departement: Life Sciences und Facility Management
Publication type: Master Thesis
DOI : 10.21256/zhaw-2059
Appears in Collections:Masterarbeiten Life Sciences

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