Abstract: This study aimed to establish a blocking ELISA method for detection of African swine fever virus(ASFV) p72 protein antibodies. With the purified recombinant p72 protein as coating antigen, and the HRP labeled 6E5 antibody as the blocking antibody, a blocking ELISA method based on ASFV p72 protein was established after optimization of the reaction conditions. One hundred and nineteen clinical negative serum samples were detected using the present established method to determine the cut off value by the serum percentage of inhibition (PI) calculation. The criteria of the blocking ELISA:serum samples with PI ≥ 50% were judged as positive; the serum samples with PI ≤ 40% were judged as negative; and the serum samples with PI between 40% and 50% were judged as suspicious. This method had no cross reactions with the serum antibodies against PRV, PRRSV, CSFV, PCV2, SVA, FMDV, Escherichia coil, Pasteurella multocida, Glaesserrella parasuis and Actinobacillus pleuropeumoniae. The intra-assay and inter-assay variation coefficients were both less than 15%. Total 447 swine clinical serum samples were then detected simultaneously by the present established method and commercial ASFV antibody detection kit. The result showed that the relative sensitivity and specificity of the two methods were 95.3% and 94.5%, respectively, and the coincidence rate was 94.9%. In summary, the blocking ELISA method established in this study has high sensitivity and specificity, therefore it is suitable for ASFV diagnosis and epidemiological investigation.

Key words: African swine fever virus, p72 protein, monoclonal antibody, blocking ELISA