Pre-colored Prussian blue: A substrate-free dual-mode immunosensor for visible-to-thermal Staphylococcus aureus quantification.

Rapid identification of postoperative Staphylococcus aureus (S. aureus) infections is essential for timely treatment and improved clinical outcomes. Herein, we developed a substrate-free dual-mode lateral flow immunoassay (LFIA) using intrinsically colored Prussian Blue nanoparticles (PB NPs) as multifunctional signal reporters. The intrinsic blue color of PB NPs enables direct colorimetric visualization, while their excellent near-infrared photothermal conversion provides quantitative temperature readout. This self-signaling strategy eliminates enzymatic amplification and substrate incubation, simplifying the assay into a one-step sandwich immunoreaction. Under optimized conditions, the proposed LFIA achieved a wide linear detection range of 10-106 CFU/mL, with detection limits of 10 CFU/mL for the colorimetric mode and 1 CFU/mL for the photothermal mode. Analysis of real samples demonstrated good agreement with a commercial ELISA kit, with relative deviations below 10% and coefficients of variation below 15.5%, confirming the accuracy and practicality of the proposed platform. The dual-readout strategy improves analytical reliability through mutual signal verification, providing a simple, rapid, and reliable approach for S. aureus detection with strong potential for point-of-care testing.