Deciphering gold nanotag-induced electron-transfer blockage in competitive electrochemical immunosensor: toward a nanoparticle-sandwiched structure for sensitive SARS-CoV-2 antigen detection.

Rapid and sensitive detection of SARS-CoV-2 remains essential for infection control and pandemic preparedness, especially with the continuous emergence of new variants. In this study, a competitive electrochemical immunosensor was developed for the detection of the S-protein using a polyethyleneimine-coated gold nanoparticle (PEI-AuNP)-based strategy. The screen-printed carbon electrode (SPCE) was modified with PEI-AuNPs to enhance surface area, conductivity, and biocompatibility, allowing efficient immobilization of the S-protein. Simultaneously, PEI-AuNPs conjugated with anti-S-protein antibodies (anti-S-protein@PEI-AuNPs) were used as nanotags. In the presence of the target antigen, antigen-antibody complexes form in the solution, reducing nanotag binding to the electrode. In contrast, the absence of the target leads to a higher density of nanotags on the electrode surface, which hinders electron transfer of the redox probe [Fe(CN)], resulting in a lower current response. The sensor demonstrated a wide linear detection range from 0.10 to 25 ng mL, with a low detection limit of 0.050 ng mL. It exhibited excellent sensitivity, selectivity, and stability. Successful detection in human serum, indoor surface swabs, and tap water with high recovery highlights its real-world applicability. This platform can be adapted for emerging pathogens, making it a valuable tool for current and future pandemic surveillance.