Glucose-fueled and wash-free immunoassay platform employing biogenic magnetic nanoparticle displaying antibody (bioMagbody).

Wash-free immunoassays with enzyme-linked immunosorbent assay (ELISA)-level performance remain challenging because peroxidase-based signal generation typically requires multiple washing steps and the handling of unstable H2O2. Here, we report a bead-based and glucose-fueled ELISA-like platform that co-localizes target capture and catalytic amplification on biogenic magnetic nanoparticles (MNPs) with peroxidase-like activity from magnetotactic bacteria. To generate biogenic antibody-displaying magnetic nanoparticle (bioMagbody), we used the SpyCatcher/SpyTag (SC/ST) system as a covalent protein coupling module. SC was genetically displayed via fusion to the abundant membrane protein Mms13 on the biogenic MNPs, enabling covalent ligation of ST-fused single-chain variable fragments (scFvs) to generate bioMagbody. SC expression and scFv display preserved the intrinsic peroxidase-like activity and target binding. Using glucose oxidase (GOx)-scFv complex as the H2O2 donor and bioMagbody as the acceptor, C-reactive protein (CRP) was quantified in buffer with a limit of detection of 4.5 nM, and specific detection was demonstrated in human serum by simple mixing without washing steps. The wash-free operation arises from target-localized H2O2 generation by GOx-scFv and proximity-enabled transfer to bioMagbody, which drives luminol chemiluminescence upon glucose addition as the reaction initiator. The platform was readily reconfigured to detect inactivated SARS-CoV-2 by exchanging scFvs. This work establishes a genetically programmable biogenic nanozyme interface for wash-free immunoassays.