Computer-aided profiling of a unique broad-specific antibody and its application to an ultrasensitive fluoroimmunoassay for five N-methyl carbamate pesticides

ABSTRACT Pollution of N-methyl carbamate (NMC) pesticides is threatening the non-target organisms’ survival. Thus, broad-specific antibodies and class-selective immunoassay are demanding for multiple NMCs determination. In this study, we employed a molecular docking-based virtual screening strategy to fast profile antibody spectrum, based on a designed chemical pool containing 17 compounds. A monoclonal antibody (mAb)-6G against carbofuran was used as the objective. The recombinant full-length IgG was successfully expressed to validate the antibody sequences for homology modeling. After docking, we manually categorized the antibody-chemical binding strength into three groups. Non-competitive surface plasmon resonance (SPR) demonstrated the mAb-6G affinitive binding toward five NMCs (carbofuran, isoprocarb, propoxur, carbaryl and carbosulfan), which were classified into strong and moderate binding categories. Antibody binding properties were confirmed again by ic-ELISA and lateral flow immunochromatographic strip. Subsequently, an ultrasensitive indirect competitive fluoromicrosphere-based immunoassay (ic-FMIA) was established with the IC50 (half-maximal inhibitory concentration) values of 0.08-3.37 ng/mL. This portable assay presented a 30~230-fold improved sensitivity than traditional ic-ELISA and was applied in European surface water analysis. Overall, our work provides an efficient platform integrating in-silico and experimental methodologies to accelerate the characterization of hapten-specific antibody binding properties and the development of high-sensitive immunoassays for multi-pollutants monitoring.