Work

COLPLEX: A high-affinity protein adaptor for enhanced sensitivity and multiplex lateral flow immunoassays. application to ebola virus strain detection

Year: 2026

Type: article

Abstract: To overcome the limitation of lateral flow immunoassays (LFIAs) in multiplexing while preserving high sensitivity and specificity, we present COLPLEX, a high-affinity recognition system based on the ultra-specific interaction between colicin antimicrobial proteins and their cognate immunity proteins. These protein pairs, naturally evolved to exhibit femtomolar binding affinities, are bio-orthogonal and readily accessible through recombinant expression. We integrated the COLPLEX system into a biplex LFIA platform for the simultaneous detection of glycoproteins from Zaire and Sudan Ebola virus variants. Using two distinct, non-cross-reactive colicin-immunity pairs, we immobilized colicins on the nitrocellulose membrane and fused immunity proteins to the capture antibodies. This configuration significantly improved the signal-to-noise ratio, reduced capture antibody consumption, and minimized non-specific binding. Clinical evaluation using sera from Ebola-infected patients (n = 30) and negative controls (n = 40), including healthy donors and patients with other viral haemorrhagic fevers, demonstrated high diagnostic accuracy, with 96.7% sensitivity and 100% specificity, confirming the robustness of COLPLEX for real-world applications. These results highlight COLPLEX as a powerful alternative to streptavidin-biotin system and its potential to revolutionize next-generation LFIA platforms. • A multiplex lateral flow immunoassay powered by COLPLEX ultra-affinity proteins delivers rapid, simultaneous, and high-sensitivity detection of Zaire and Sudan Ebola virus strains • High-affinity colicin-immunity protein interaction was harnessed • Approach outperforms classic direct immunoassays in sensitivity • Clinical evaluation confirms high sensitivity and no non-specific binding

Source: Sensors and Actuators B Chemical

DOI: 10.1016/j.snb.2026.139608