Preparation and Characterization of Polyclonal Antibodies Against Porcine Enterovirus G-PLP 3C Protein

Porcine enterovirus G (EV-G), a member of the Enterovirus genus in the Picornaviridae family, is a small RNA virus that commonly infects pigs. While often asymptomatic, EV-G can contribute to diarrhea in pigs, especially in conjunction with other gastrointestinal viruses. Recent years have seen an increase in the emergence of EV-G strains with naturally recombinant PLP genes globally. This study focuses on the development of EV-G-PLP-specific antibodies and the investigation of the virus's ability to infect mice.

Preparation of EV-G-PLP 3C Polyclonal Antibody

To generate polyclonal antibodies targeting the EV-G-PLP non-structural 3C protein, we constructed the recombinant plasmid pET-32a-EV-G-PLP-3C and transformed it into BL21 (DE3) cells. We optimized the induction conditions (temperature, IPTG concentration, and induction time) to maximize the yield of 3C recombinant protein. After purifying the 3C protein, we immunized New Zealand rabbits via subcutaneous injection to produce polyclonal antibodies. We then evaluated the antibody titer using indirect ELISA and confirmed specificity through indirect immunofluorescence assay and Western blot analysis.

Our results demonstrated that the 3C protein primarily existed as inclusion bodies with a molecular weight of approximately 38.8 kDa. The generated polyclonal antibodies exhibited strong immunogenicity and reactivity. This study offers valuable insights into diagnosing EV-G-PLP infections and understanding the biological functions of the 3C protein.