A novel one-step immunoassay using mesoporous core-shell Pd@Pt nanoparticles as an alternative to horseradish peroxidase for amantadine detection

The inherent limitations of natural enzymes hinder their stable application under harsh conditions, highlighting the urgent need for a stable alternative signal tracer. Herein, core-shell palladium@platinum (Pd@Pt) nanoparticles (NPs) with mesoporous and dendritic nanostructures were synthesized using a modified ultrasound-assisted chemical reduction method, resulting in a 25 % improvement in relative activity compared to previously reported methods. The yielding nanoparticles exhibited enhanced peroxidase-like activity with specific activity of 54.4 U/mg, and their Michaelis constant and maximum reaction velocity were 6.2 and 48 timers higher, respectively, than those of horseradish peroxidase (HRP). The Pd@Pt NPs were conjugated with antibodies via electrostatic adsorption, demonstrating their potential as a viable alternative to HRP. The rough surface structure of the Pd@Pt NPs not only enhanced the efficiency of antibody immobilization to approximately 90 %, but also, through a pH-regulated antibody immobilization strategy, facilitated the orientation of antibody recognition regions, thereby improving inhibition efficiency. A one-step immunoassay based on the Pd@Pt NPs immunoprobe was then developed, with amantadine (AMD) serving as a proof of concept. The proposed method showed a 2.8-fold increase in sensitivity, a 400-fold expansion in the linear range, and a 1-h reduction in detection time compared to the traditional “gold standard” ELISA. The practical applicability of the proposed method was validated in real chicken and pork samples, with coefficients of variation all less than 17.49 %, and a strong correlation with commercial kits was observed. These results suggest that Pd@Pt NPs are a promising signal-generating probe with significant potential as an alternative to HRP, offering improvements in sensitivity, linear range, stability, and other key performance characteristics.