Evaluating the Immunogenicity of Avian Influenza Virus Nucleoprotein
2020
Zahra Sadat Khademi Sayed Bonadaki | Rasool Madani | Parviz Pakzad | Fariba Golchinfar | Tara Emami
Background: Influenza viruses cause Avian Influenza (AI) is a serious infectious disease belonging to type A Orthomyxovirus. A viral RNA synthesis is due to an interaction of the nucleoprotein (NP) with the viral polymerase. In the present study, we have evaluated the immunogenicity of avian influenza virus nucleoprotein. Materials & Methods: An Influenza Virus N9H2 subtype A/Chicken Iran/259/2014 was selected. In order to perform electrophoresis and purification of nucleoprotein, the protein concentration of the samples was determined. SDS-PAGE and Native-PAGE electrophoresis on polyacrylamide gel were done and differences in gel bands in two methods were compared. Purification of the virus nucleoprotein performed by electroelution method and purified nucleoprotein were assayed by ELISA to obtain the produced antibody titers. Ouchterlony and western blot tests were used for final approval.Results: By determining the molecular weight of each polypeptides, the molecular weights of H9N2 proteins were ranged from 30 to 140kDa and the molecular weight of the nucleoprotein was 60kDa. The nucleoprotein was purified by electroelution method. Ouchterlony showed that in serum dilution of 1:2 and 1:4, the sediment lines were formed between the serum and the NP antigen. The NP-ELISA enables rapid serological diagnosis in 1:20. Finally, the western blot test confirmed the 60kDa nucleoprotein band.Conclusion: The nucleoprotein which was purified by electroelution retained its antigenic property and it could be applied in diagnostic kits.
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