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Selection of Affibody Molecules Using Staphylococcal Display
KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Protein Engineering.ORCID iD: 0000-0001-9423-0541
KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science.ORCID iD: 0000-0001-6558-0702
KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Protein Technology.ORCID iD: 0000-0002-9952-9814
KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Protein Technology.ORCID iD: 0000-0002-9282-0174
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2024 (English)In: Cold Spring Harbor Protocols, ISSN 1940-3402, E-ISSN 1559-6095, Vol. 2024, no 11Article in journal (Refereed) Published
Abstract [en]

Affibody molecules are small (6-kDa) affinity proteins generated by directed evolution for specific binding to various target molecules. The first step in this workflow involves the generation of an affibody library, which can then be used for biopanning using multiple display methods. This protocol describes selection from affibody libraries using display on Staphylococcus carnosus. Display of affibodies on staphylococci is very efficient and straightforward because of the single cell membrane and the use of a construct with a constitutive promoter. The workflow involves display of affibody libraries on the surface of S. carnosus cells, followed by screening and selection of binders using fluorescence-activated cell sorting (FACS). The transformation of DNA libraries into S. carnosus is less efficient and more complicated than for Escherichia coli. Because of this, staphylococcal display is suitable for affinity maturation or other protein-engineering efforts that are not dependent on very high diversity, and thus magnetic-activated cell sorting (MACS) is often not required before FACS. However, MACS is an option, and MACS procedures used for E. coli can easily be adapted for use in S. carnosus if needed.

Place, publisher, year, edition, pages
Cold Spring Harbor Laboratory , 2024. Vol. 2024, no 11
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Molecular Biology
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URN: urn:nbn:se:kth:diva-366513DOI: 10.1101/pdb.prot108401PubMedID: 37491081Scopus ID: 2-s2.0-85208289045OAI: oai:DiVA.org:kth-366513DiVA, id: diva2:1982575
Note

QC 20250708

Available from: 2025-07-08 Created: 2025-07-08 Last updated: 2026-03-20Bibliographically approved

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Löfblom, JohnHjelm, Linnea C.Dahlsson Leitao, CharlesStåhl, StefanLindberg, Hanna

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