Preparation Of Specific Antiserum Against Structure Protein Of Newcastle Disease Virus

Newcastle disease(ND), which could infect wild range of birds, including chicken, turkey and 200 other species of birds, is caused by Newcastle Disease Virus. ND outbreak often results in severe economic losses because of high morbidity and mortality. It’s listed as a notifiable disease by Office International Des Epizooties(OIE). Although the outbreak and prevalence of ND has been controlled in China, the latent infection and prevalence in vaccinated chickens still exist. Improving technology of detection and developing new immunological evaluation methods become the focus of epidemic prevention and control.Newcastle disease virus includes six structure proteins,they are large polymerase protein(L), hemagglutinin-neuraminidase protein(HN), fusion protein(F), nucleocapsid protein(NP), phosphoprotein( P) and matrix protein(M). In this study, the viral protein was separated by SDS-PAGE. And then guinea pigs were immunized with these proteins recovery. Finally, HN protein antiserum and NP antiserum were prepared successfully.1. Purification of virions. The 30% sucrose ultracentrifugation, which needs only one density gradient, was prove to be a very simple method for virus purification. The HA titer of NDV could reach 15log2 and the viral concentration was 5.3 mg/mL. In addition, the purity of viral was about 99.4%.2. Isolation and preparation of NDV proteins. We investigated the protein preparation method based on SDS-PAGE. After SDS-PAGE, the polyacrylamide gel was stained with 0.25 M KCl. Then the gel contain different protein was excised, and rinsed with distilled water. Furthermore, the protein in gel was recovered with solutions that SDS was main components.3. Animal experiment. The HN protein and NP protein prepared by gel extraction could elicit a strong immune response in guinea pigs. And the HN protein antiserum and NP antiserum were prepared successfully. Through identification, HN protein antiserum showed good specificity, and the antibody titers was 20,000. The NP antiserum antibody titer was about 60,000.