Zein–templated synthesis of Au@Pt bimetallic nanozymes: a multi–mode lateral flow immunoassay platform for ultrasensitive colorimetric and fluorometric detection of tetracycline in food

Abstract

The systematic development of a multifunctional zein–stabilized Au@Pt bimetallic nanozyme (zein–Au@Pt) for highly efficient multimodal detection of tetracycline (TC) is described. We created a core-shell nanostructure using zein protein as a biomimetic template, combining the Au@Pt bimetallic interface’s high peroxidase-mimicking activity with the prolamine shell’s fluorescence. Zein–Au@Pt exhibited low Michaelis constants (𝐾𝑚 for H₂O₂: 0.813 mM; TMB: 0.211 mM) in steady-state kinetics, indicating high substrate affinity. A multimodal lateral flow immunoassay (LFIA) platform allows traditional colorimetric screening, catalytically amplified detection, and fluorescence measurement. These three approaches had detection limits of 0.9, 0.09, and 0.014 ng/mL, respectively, demonstrating 10-fold (catalytic amplification) and 64-fold (fluorescence sensitization) increases in sensitivity over traditional colorimetric LFIA. High recoveries (89.8–109.3%) across various complex animal-derived matrices demonstrated its adaptability as a multimodal LFIA platform for on-site screening after minimal sample preprocessing for antibiotic residue surveillance. Despite these benefits, challenges remain, including potential cross-reactivity with TC analogs in complex matrices, the need to miniaturize the fluorescence readout device for true point-of-care use, and the long-term stability of the zein protein coating under different storage conditions. Engineering highly specialized recognition elements (aptamers) to overcome cross-reactivity, integrating smartphone-based detecting systems for mobility, and scaling up zein-templated nanozymes for commercial use are future priorities.

Graphical abstract