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Discontinuous epitope mapping of antibodies using bacterial cell surface display of folded domains
KTH, School of Biotechnology (BIO).ORCID iD: 0000-0003-1096-9061
KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Protein Technology.ORCID iD: 0000-0002-9977-5724
2018 (English)In: Epitope Mapping Protocols, Humana Press, 2018, p. 159-183Chapter in book (Refereed)
Abstract [en]

Knowledge of the exquisite-binding surface of an antibody on its target protein is of great value, in particular for therapeutic antibodies for understanding method of action and for stratification of patients carrying the necessary epitope for desired drug efficacy, but also for capture assays under native conditions. Several epitope mapping methodologies have been described for this purpose, with the laborious X-ray crystallography method being the ideal method for mapping of discontinuous epitopes in antibody-antigen crystal complexes and high-throughput peptide-based methods for mapping of linear epitopes. We here report on the usage of a bacterial surface display-based method for mapping of structural epitopes by display of folded domains on the surface of Gram positive bacteria, followed by domain-targeted mutagenesis and library analysis for the identification of key-residues by flow sorting and sequencing. Identified clones with reduced affinity are validated by single clone FACS and subsequent full-length expression in mammalian cells for validation.

Place, publisher, year, edition, pages
Humana Press, 2018. p. 159-183
Series
Methods in Molecular Biology, ISSN 1064-3745 ; 1785
Keywords [en]
Antibody, Cell surface display, CHO, Discontinuous epitope mapping, FACS, Gram-positive, S. carnosus
National Category
Medical Biotechnology
Identifiers
URN: urn:nbn:se:kth:diva-238351DOI: 10.1007/978-1-4939-7841-0_11Scopus ID: 2-s2.0-85046359816ISBN: 978-1-4939-7839-7 (print)OAI: oai:DiVA.org:kth-238351DiVA, id: diva2:1262805
Funder
Knut and Alice Wallenberg Foundation
Note

QC 20181113

Available from: 2018-11-13 Created: 2018-11-13 Last updated: 2018-11-13Bibliographically approved

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Volk, Anna-LuisaRockberg, Johan

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CiteExportLink to record
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