LAUSR

BACKGROUND African swine fever virus (ASFV) is the etiologic agent of African swine fever (ASF), a disease of highly contagious and significant threat to pork production. At present, the sensitive detection methods are the keys to the disease control. OBJECTIVE Full length p72 is produced by eukaryotic system and its monoclonal antibody (mAb) 34C10 is subsequently recovered. A blocking ELISA kit for detection of ASFV antibody is developed based on p72 trimers and 34C10. METHODS Full length p72 is expressed and is used as immunogen to prepare a panel of monoclonal antibodies. The mAb 34C10 is verified by immunofluorescent and tested by ELISAs with positive serums. The constant affinity of 34C10 is then confirmed. A blocking ELISA kit is further developed and is compared with two commercial kits. RESULTS The mAb 34C10 is specifically bound to p72 protein, and it exhibits blocking affection to positive serum. IFA experiment shows that 34C10 could bind to p72 expressed by baculoviruses and the binding affinity of 34C10 is found to be as high as 1.85 × 1011 L/mol. The blocking ELISA kit shows high coincidence with a commercial ELISA kit. The sensitivity between these two kits is 97.6% (95%, CI: 90.65-99.58) and the specificity between them is 100% (95%, CI: 98.34-100). CONCLUSION The blocking ELISA developed in this study may have great potential for diagnosis of ASF. The structure of the antigen p72 is found to be a key factor for the performance of the kit. HIGHLIGHTS For the first time, the eukaryotic expressed full-length p72 protein is utilized to recover the monoclonal antibody and it is coated as antigen during the development of the blocking ELISA kit. This study sheds new light on the development of the blocking ELISA kits, especially for the development of diagnostic kit for the contagious virus with bio-safety problems.