Oriented antibody coupling enhances lateral flow immunoassay sensitivity for imidacloprid detection in vegetables and fruits

Lateral flow immunoassay (LFIA) offers advantages for rapid, on-site detection of pesticides. Herein, aggregation-induced emission fluorescent microspheres were modified using maleimide groups to enable site-specific coupling of recombinant protein A (rPA). Subsequently, the Fc region of an anti-imidacloprid monoclonal antibody was bound to rPA to create an oriented immunoprobe. LFIA with the oriented immunoprobe was developed for detecting imidacloprid, a widely misused neonicotinoid pesticide. The Fab orientation ratio of the oriented immunoprobe reached 91.1%, when compared with 58.1% for the non-oriented immunoprobe. The oriented immunoprobe with a higher orientation improved immunobinding kinetics, enhanced the LFIA signal, and increased assay sensitivity. The limit of detection was 0.12   ng/mL, which was lower than 0.31   ng/mL that was achieved with the non-oriented immunoprobe. In addition, LFIA with the oriented immunoprobe required 40% less antibody and 25% less coating antigen. The recovery results were consistent with HPLC–MS/MS, demonstrating accurate detection of imidacloprid in cabbage, chili, and orange. The LFIA with oriented immunoprobe proved to be a rapid and sensitive method for food-safety monitoring.