Development of functionalized gold nanoparticles as nanoflare probes for rapid detection of classical swine fever virus

Abstract Classical swine fever (CSF) is a devastating viral disease affecting pigs that causes major economic losses worldwide. Conventional assays to identify classical swine fever virus (CSFV) face challenges, such as the required molecular amplification of the target molecules via polymerase chain reaction (PCR). We designed a gold nanoflare probe to directly detect CSFV. Gold nanoparticles (AuNPs) were conjugated with a pair of complementary DNA sequences that specifically recognized and captured CSFV RNA, resulting in a fluorescence signal to indicate the existence of CSFV. The constructed nanocomposite was then utilized in a quantitative analysis to recognize the virus sequence present at amounts as low as 50 pg/μL. The CSFV-AuNP probe enabled real-time, quantitative detection of native CSFV in response to doses of the specific RNA sequence (CSFV NS2) that indicated active viral replication of CSFV Shimen in macrophages after 12, 24, and 48 h. The potential diagnostic applications of the probe were demonstrated by measuring CSFV without nucleic acid amplification in samples from seven types of tissue samples, specifically heart, spleen, kidney, liver, lymph, intestine, and muscle samples obtained from one pig confirmed to suffer CSF. The speed, sensitivity, and versatility of this CSFV-AuNP biosensor make it an ideal candidate for further application in the prevention and control of animal epidemic diseases.