Ta DT Vanella R Nash MA 2018 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. application/pdf https://sonar.ch/global/documents/15540 eng info:eu-repo/semantics/altIdentifier/doi/10.1021/acsami.8b10092 info:eu-repo/semantics/altIdentifier/pmid/30125079 info:eu-repo/semantics/openAccess ACS applied materials & interfaces. - 2018 ELP SPAAC multiprotein complex protein polymer single-domain antibody Cycloaddition Reaction Elastin Humans Immunoassay Limit of Detection Luminescent Proteins Peptides Bioorthogonal Elastin-like Polypeptide Scaffolds for Immunoassay Enhancement. http://purl.org/coar/resource_type/c_6501