Abstract: To explore the B cell epitopes of African swine fever virus (ASFV) p30 protein, the monoclonal antibody (mAb) of p30 protein was prepared, and it was used as a tool for screening its B cell epitopes. First of all, p30 protein was obtained by prokaryotic expression system and purified by Ni affinity method. BALB/c mice were immunized with purified protein and hybridoma cells were prepared. Positive hybridoma cells were screened by indirect enzyme linked immunosorbent assay (iELISA). The mAb was then prepared by cell expression method. The specificity of mAb was identified by indirect immunofluorescence assay (IFA) and Western blot. Then, IEDB epitope prediction software was used to predict the B cell epitopes of p30 protein. According to the prediction results, CP204L gene was truncated to express in prokaryotic cells. IFA, Western blot and iELISA were used to identify the epitopes. Finally, a phage-displayed 12-mer peptide library was used to conduct four rounds of biological panning for the p30 monoclonal antibody, and identify the specific epitope of p30, which was further compared with the truncated protein mapping described above. The results of specific identification showed that the mAb could successfully identify ASFV in porcine alveolar macrophages. The results of truncated protein mapping showed that the epitopes region recognized by the mAb of p30 protein was ⁸⁴M-K¹⁴². The biopanning experiment indicated that ¹¹⁶TSSFETLFE¹²⁴ was a core domain of the B cell linear epitope of p30 protein, which further narrowed the range of the epitope distribution. Altogether, a mAb against ASFV p30 protein was prepared, and the epitopes were identified, which provided a reference for the development of serological diagnostic reagents and lay a foundation for the study of p30 protein function

Key words: African swine fever virus, p30 protein, monoclonal antibody, biopanning, epitope