A Host–Guest-Regulated Photocurrent Polarity Switching with a “Binary” Strategy for False-Positive-Free Microbial Detection of <i>Staphylococcus aureus</i>

triggers a unique photocurrent polarity switch from anodic to cathodic. Mechanistic studies reveal that this inversion stems from the encapsulation of the terminal alkyne group of TEPy by CB [7], which alters the electron transfer pathway between the complex and the bacteria. This SPI mechanism not only amplifies the detection signal but also effectively suppresses false-positive responses in complex matrices through polarity switching. Under optimized conditions, the biosensor exhibits excellent selectivity over competing bacteria and achieves a remarkably low detection limit of 2.76 CFU/mL. It also demonstrates high applicability in real food samples. Furthermore, molecular docking simulations provide insights into the assembly and sensing mechanisms. This work significantly mitigates interference issues inherent in conventional detection methods, offering a rapid, efficient, and accurate analytical platform for microbial detection in food and environmental monitoring.