Development of Au@SiO2 nanostructure-based metal-enhanced fluorescence biosensor for African swine fever virus antibody detection.

African swine fever (ASF) is a devastating viral disease of domestic pigs and wild boar that continues to cause substantial economic losses to the global swine industry. Although molecular methods such as PCR remain the reference standard for early detection of acute infection, serological assays play an important role in antibody surveillance and herd management. In this study, we developed a metal-enhanced fluorescence (MEF) biosensor for the highly sensitive detection of African swine fever virus (ASFV)-specific antibodies. The sensor platform consists of a gold nanoparticle (AuNP) array fabricated by DC magnetron sputtering and thermal dewetting, followed by the deposition of an optimized silica (SiO2) spacer layer using RF sputtering. The resulting Au@SiO2 substrate exhibited a localized surface plasmon resonance (LSPR) peak at approximately 565 nm, providing efficient spectral overlap with the Cy3 fluorophore. Surface characterization by water contact angle (WCA) measurements confirmed successful stepwise biofunctionalization from amine grafting to antigen immobilization and sequential antibody binding. The developed MEF platform demonstrated high analytical sensitivity, detecting ASFV antibodies at dilutions up to 1:10,240 with a clear concentration-dependent fluorescence response and high reproducibility (%RSD = 9.45%) across three fabrication batches. In addition, preliminary evaluation using ASFV-positive swine serum demonstrated the feasibility of the platform for serological antibody detection. These findings suggest that the proposed MEF biosensor represents a promising high-sensitivity platform for ASFV serological screening and antibody monitoring, complementing existing molecular diagnostic methods.