Show simple item record

dc.contributor.authorFeng, M.
dc.contributor.authorBian, H.
dc.contributor.authorFlajnik, M.F.
dc.date.accessioned2020-03-30T17:40:05Z
dc.date.available2020-03-30T17:40:05Z
dc.date.issued2018
dc.identifier.urihttps://www.scopus.com/inward/record.uri?eid=2-s2.0-85071516744&doi=10.1093%2fabt%2ftby011&partnerID=40&md5=03a0372921bddd2f01acf64ba422e5d7
dc.identifier.urihttp://hdl.handle.net/10713/12445
dc.description.abstractBackground: Shark new antigen receptor variable domain (VNAR) antibodies can bind restricted epitopes that may be inaccessible to conventional antibodies. Methods: Here, we developed a library construction method based on polymerase chain reaction (PCR)Extension Assembly and Self-Ligation (named “EASeL”) to construct a large VNARantibody library with a size of 1.2 × 1010from six naïve adult nurse sharks (Ginglymostoma cirratum). Results: The next-generation sequencing analysis of 1.19 million full-length VNARs revealed that this library is highly diversified because it covers all four classical VNARtypes (Types I–IV) including 11% of classical Type I and 57% of classical Type II. About 30% of the total VNARs could not be categorized as any of the classical types. The high variability of complementarity determining region (CDR) 3 length and cysteine numbers are important for the diversity of VNARs. To validate the use of the shark VNARlibrary for antibody discovery, we isolated a panel of VNARphage binders to cancer therapy-related antigens, including glypican-3, human epidermal growth factor receptor 2 (HER2), and programmed cell death-1 (PD1). Additionally, we identified binders to viral antigens that included the Middle East respiratory syndrome (MERS) and severe acute respiratory syndrome (SARS) spike proteins. The isolated shark single-domain antibodies including Type I and Type II VNARs were produced in Escherichia coli and validated for their antigen binding. A Type II VNAR(PE38-B6) has a high affinity (Kd= 10.1 nM) for its antigen. Conclusions: The naïve nurse shark VNARlibrary is a useful source for isolating single-domain antibodies to a wide range of antigens. The EASeL method may be applicable to the construction of other large diversity gene expression libraries. Copyright The Author(s) 2018.en_US
dc.description.urihttps://doi.org/10.1093/abt/tby011en_US
dc.language.isoen_USen_US
dc.publisherOxford University Pressen_US
dc.relation.ispartofAntibody Therapeutics
dc.subjectGene library constructionen_US
dc.subjectNext-generation sequencingen_US
dc.subjectPhage displayen_US
dc.subjectShark VNARen_US
dc.subjectSingle-domain antibodyen_US
dc.subject.meshMiddle East Respiratory Syndrome Coronavirusen_US
dc.subject.meshSARS Virusen_US
dc.titleConstruction and next-generation sequencing analysis of a large phage-displayed VNARsingle-domain antibody library from six naïve nurse sharksen_US
dc.typeArticleen_US
dc.identifier.doi10.1093/abt/tby011
dc.identifier.ispublishedNo


This item appears in the following Collection(s)

Show simple item record