Identification of Linear B Cell Epitopes on CD2V Protein of African Swine Fever Virus by Monoclonal Antibodies

The CD2-like (CD2V) protein is a crucial antigen of African swine fever virus (ASFV). CD2V interacts with the cellular AP-1 protein, participates in intracellular transport of virus, and induces neutralizing antibodies to partly protect swine from virus attack. In this study, a specific CD2V dimeric protein was designed to enhance antigenicity and immunogenicity, expressed in a Bac-to-Bac baculovirus expression vector system and purified by Ni-affinity chromatography. After animal immunization, five monoclonal antibodies (mAbs) (7E12, 22B3, 18A3, 13G11, and 43C2) against CD2V were developed. The variable regions of heavy chains and light chains of the mAbs were sequenced to prove that the five mAbs differed from one another. The mAbs of CD2V could combine with ASFV by immunoperoxidase monolayer assay (IPMA). B cell epitopes of CD2V were screened using the five mAbs by indirect enzyme-linked immunosorbent assay (ELISA) and Dot-ELISA. Therefore, three B cell epitopes ((147)FVKYT(151), (157)EYNWN(161), and (195)SSNY(198)) were identified. This is the first time that mAbs of the ASFV CD2V protein have been developed and the sequencing of heavy chains and light chains of mAbs has been completed. Linear B cell epitopes, which were core targets of immunoprotection of the CD2V protein, were identified by mAbs for the first time. This study provides efficient epitopes for the development of ASFV subunit vaccines. IMPORTANCE The ASFV CD2V protein is a crucial antigen on the outer envelopes of virus particles. A modified ASFV CD2V dimeric protein was expressed in the Bac-to-Bac baculovirus expression vector system. Five monoclonal antibodies (mAbs) against CD2V were developed, sequenced, and applied to identify CD2V protein B cell epitopes. Three B cell epitopes, (147)FVKYT(151), (157)EYNWN(161), and (195)SSNY(198), were identified. This is the first time CD2V mAbs have been developed, the sequencing of heavy chains and light chains of CD2V mAbs have been completed, and CD2V B cell epitopes have been identified by using scanning peptide method and bioinformatics methods.