Establishment of an indirect ELISA method and development of corresponding kit targeting African swine fever virus p54 antibodies
The African swine fever virus(ASFV)is a highly contagious viral pathogen that poses a significant threat to the global pig industry.So far,no effective vaccine is available for the disease.The p54 protein,a prominent structural component of ASFV,is involved in virus adsorption and subsequent entry into cells during the invasion process.The protein also assumes a pivotal role in diagnostic techniques.In the present study,an optimized indirect enzyme-linked immunosorbent as-say(ELISA)was successfully developed,employing the non-tagged p54 protein as coating anti-gens.The cutoff value of the kit was determined by evaluating 374 pig ASFV negative serum sam-ples and preparation of the serum standard for the quality control of the kit.Furthermore,the sen-sitivity,specificity and repeatability of different pig pathogen positive quality control sera were e-valuated.The results showed that the established cutoff value of the kit was 0.250.The detection limit of the kit for control positive serum was up to a dilution ratio of 1:1 600.No cross-reactivity was detected for the antibodies to CSFV,PRRSV,PCV2,PRV,PPV,or E.coli Rosetta.Both the coefficients of intra-and inter-assay variability were found to be below 10％.The specificity and sensitivity remained unchanged when the kit was stored at 2-8 ℃ for 9 months.In conclusion,this antibody kit for ASFV exhibits a notable level of sensitivity,specificity,and stability,and has a high compliance rate with the OIE-recommended p72 blocking ELISA antibody detection kit,which provides strong technical support for the detection of ASFV antibodies and serological sur-veillance for African swine fever.
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