Development of Visual Detection of African Swine Fever Virus Using CRISPR/AapCas12b Lateral Flow Strip Based on Viral...

African swine fever (ASF), induced by the African swine fever virus (ASFV), is an acute hemorrhagic disease characterized by high fever, systemic hemorrhages, and elevated mortality. Current diagnostic techniques including PCR and ELISA present limitations in field applications due to requirements for specialized equipment and prolonged processing duration. Therefore, rapid and accurate detection of ASFV has become a key link in ASF prevention and control. This study established a rapid and precise visual diagnostic approach by integrating the CRISPR/AapCas12b system with lateral flow strip (LFS) technology, specifically targeting the B646L gene encoding the major capsid protein p72. The CRISPR/AapCas12b-LFS platform achieved a sensitivity threshold of 6 copies/µL for B646L gene detection, completing analysis within an hour. Validation study confirmed exceptional specificity against common porcine pathogens including PRRSV, CSFV, PRV, PPV4, and PCV3. The developed assay demonstrated complete concordance with real-time PCR results when analyzing 34 clinical specimens including three heart samples, three liver samples, three spleen samples, three lung samples, three kidney samples, three lymph node samples, five serum samples, five blood samples, and five oral swab samples for ASFV detection. Overall, this method is sensitive, specific, and practicable onsite for ASFV detection, showing a great application potential for monitoring ASFV in the field.