Preparation of monoclonal antibody against porcine saperovirus VP1 protein and identification of its antigenic epitope

ABSTRACT

As a member of the family Picornaviridae, porcine sapelovirus (PSV) is often infected with porcine epidemic diarrhea virus, teschovirus and so on. In recent years, PSV has been isolated from porcine in many provinces of China. It suggests that it is necessary to strengthen the research on PSV. In this study, according to the sequence of PSV HuN2 strain, VP1 gene was inserted into the pGEX-6 P-1 vector, and expressed the recombinant protein. BALB/c mice aged 6-8 weeks were immunized according to the standard procedure. After the third immunization, the mouse orbital blood was collected to identify the antibody level. The highly positive mouse spleen cells were selected for cell fusion. The positive hybridoma cells and two subclones were screened by IFA method, and then a PSV VP1 monoclonal antibody was obtained, named as 33-2 A. The results of IFA showed that PSV could be recognized by 33-2 A MAb, and specific green fluorescence appeared in the cytoplasm;The results of WB and IP showed that PSV infected porcine cell could specifically bind to 33-2 A, and there was a specific band at 32 ku. We also identified the B-cell antigen epitope of 33-2 A, it was at amino acids 40-46 of PSV VP1 protein, and the polypeptide sequence was 40PALTAAE46. The results showed that the monoclonal antibody can react with PSV VP1 protein. The epitope was analyzed with the PSV sequences uploaded in NCBI, 33-2 A antibody can react with most PSV strains and has a certain universal to PSV. This study laid a foundation for the study of the etiology and pathogenesis of PSV.