A ultrasensitive immunoassay based on weakly charged AuNPs orientated antibody for triazophos detection in vegetables

Immunoassays based on gold nanoparticles (AuNPs) often suffer from limited sensitivity. Herein, we synthesized weakly charged AuNPs using ascorbic acid and establish a competitive immunoassay for the detection of triazophos. The weak electronegativity of these AuNPs reduces electrostatic repulsion during antibody adsorption, leading to increased antibody loading and favorable antibody orientation, thereby enhancing detection sensitivity. The triazophos antibody and horseradish peroxidase (HRP) were electrostatically adsorbed onto the AuNP surface to form an antibody-AuNP-enzyme probe complex. In the immunoassay, triazophos in the sample competes with the coated antigen for binding to the Ab@AuNPs@HRP probe, and quantification is achieved through an HRP-catalyzed colorimetric reaction. The proposed method demonstrated a linear range of 1.08-22.48 µg/L, an IC₅₀ value of 4.93 µg/L, and a detection limit as low as 0.06 µg/L. This approach eliminates the need for enzyme-labeled secondary antibodies, offering advantages such as lower cost, simpler operation, higher sensitivity, and good specificity.