Multimodal Lateral Flow Immunoassay Based on Four-in-One Au@Prussian Blue Nanoparticles for Ultrasensitive Higenamine Screening in Urine and Food Samples

Higenamine (HIG) has been officially classified as a sports-doping agent by the World Antidoping Agency (WADA). Considering its wide presence in plants and food items, the establishment of rapid, sensitive, and on-site detection methods for HIG is extremely crucial. Such assays can effectively assist athletes in preventing unintentional ingestion of HIG. In this study, a multifunctional nanoprobe, named Au@Prussian blue (Au@PB NPs), was synthesized. These Au@PB NPs not only generate a visible color signal but also possess a surface-enhanced Raman scattering (SERS) property, photothermal effect, and peroxidase-like (POD) activity. These characteristics enable multimode output signals, including visual color, Raman, temperature, and colorimetric signals. With the aid of HIG antibodies, this “four-in-one” nanoprobe was employed to develop a multimodal lateral flow immunoassay (LFIA) for detecting HIG in urine and food samples. Notably, the unique Raman signal of Au@PB within the Raman-silent region (1800–2800 cm–1) ensures a high signal-to-background ratio. As a result, an ultralow limit of detection (LOD) for HIG, down to 0.01 ng/mL, can be achieved, which is significantly lower than most of the existing methods. Moreover, the other output models can help minimize false positives or negatives through multisignal verification and enhance application flexibility by using different output signals and adapting to various concentration ranges. Therefore, this assay offers a rapid, sensitive, selective, and on-site detection approach for HIG, providing a powerful tool for the prevention and detection of stimulants.