Bioorthogonal Elastin-like Polypeptide Scaffolds for Immunoassay Enhancement

Ta, Duy Tien and Vanella, Rosario and Nash, Michael A.. (2018) Bioorthogonal Elastin-like Polypeptide Scaffolds for Immunoassay Enhancement. ACS Applied Materials and Interfaces, 10 (36). pp. 30147-30154.

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Official URL: https://edoc.unibas.ch/67938/

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Artificial multiprotein complexes are sought after reagents for biomolecular engineering. A current limiting factor is the paucity of molecular scaffolds which allow for site-specific multicomponent assembly. Here, we address this limitation by synthesizing bioorthogonal elastin-like polypeptide (ELP) scaffolds containing periodic noncanonical l-azidohomoalanine amino acids in the guest residue position. The nine azide ELP guest residues served as conjugation sites for site-specific modification with dibenzocyclooctyne (DBCO)-functionalized single-domain antibodies (SdAbs) through strain-promoted alkyne-azide cycloaddition (SPAAC). Sortase A and ybbR tags at the C- and N-termini of the ELP scaffold provided two additional sites for derivatization with small molecules and peptides by Sortase A and 4`-phosphopantetheinyl transferase (Sfp), respectively. These functional groups are chemically bioorthogonal, mutually compatible, and highly efficient, thereby enabling synthesis of multi-antibody ELP complexes in a one-pot reaction. We demonstrate application of this material for enhancing the performance of sandwich immunoassays of the recombinant protein mCherry. In undiluted human plasma, surfaces modified with multi-antibody ELP complexes showed between 2.3- and 14.3-fold improvement in sensitivity and ∼30-40% lower limits of detection as compared with nonspecifically adsorbed antibodies. Dual-labeled multi-antibody ELP complexes were further used for cytometric labeling and analysis of live eukaryotic cells. These results demonstrate how multiple antibodies complexed onto bioorthogonal protein-based polymers can be used to enhance immunospecific binding interactions through multivalency effects.
Faculties and Departments:05 Faculty of Science > Departement Chemie > Chemie > Synthetic Systems (Nash)
UniBasel Contributors:Nash, Michael and Vanella, Rosario and Ta, Duy Tien
Item Type:Article, refereed
Article Subtype:Research Article
Publisher:American Chemical Society
Note:Publication type according to Uni Basel Research Database: Journal article
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edoc DOI:
Last Modified:17 Oct 2019 07:41
Deposited On:06 Feb 2019 14:23

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