Study On The Immunogenicity Of Porcine Parvovirus Capsid Protein Carrying Neutralizing Epitope Of Foot-and-Mouth Disease Virus

Foot and mouth disease on China's aquaculture industry poses a serious threat,due to the traditional inactivated vaccine virus virulence back,inactivation is not complete and other unsafe factors,prompted people to study a more safe foot and mouth disease vaccine.PPV-VLPs subunit vaccine has similar stability and immunogenicity of natural viruses,does not have viral nucleic acid,no infectivity,and therefore has a good application prospects.In this study,the recombinant virus was used as the material to study the recombinant virus and the optimal expression condition and immunogenicity of the recombinant protein were studied by using baculovirus expression system.In order to develop a cheap,efficient and safe PPV-VLPs subunit vaccine.In this paper,the optimal culture mode of insect cells was studied by comparing the cell morphology of insect cell suspension culture and adherent culture.Cell counting was performed every 24 h after counting with blood cell count plate.The morphological curves of insect cells were studied.In order to obtain high titer of P3 recombinant virus,the P1 recombinant virus was transfected into insect cells,and the viral DNA was extracted,PCR was used to identify whether the inserted foreign gene was lost.Western blotting assay was used to detect the expression of recombinant protein and its reactivity.Using different MOI infected insect cells,different time to harvest the virus,baculovirus titration kit to determine the virus titer,the study of recombinant virus value-added process.The cells were harvested with different MOI,and the cells were harvested at different time.Western-blotting analysis was performed.The optimal expression conditions of recombinant protein were studied by software.A large number of suspended insect cells were infected with the best recombinant protein expression conditions.The recombinant protein was expressed in a large amount.The crude protein expression product was emulsified and injected into guinea pigs.The serum was collected and the antibody level was detected by indirect ELISA.The results showed that:(1)the optimal culture mode of insect cells was suspension culture;(2)In order to obtain high titer of P3 recombinant virus,PCR identification of inserted foreign gene is not lost because of passage.(3)To study the recombinant virus value-added process,the optimal MOI was 0.05 MOI,the best infection time was 72h;(4)The recombinant protein was in the presence of the recombinant protein,The results showed that the reactivity of the recombinant protein was good,and the presence of the neutralizing epitope of the foot-andmouth disease virus was further confirmed.(5)The optimal expression conditions of therecombinant protein were studied by Western-blotting The results showed that the optimal conditions were 10 MOI,the best infection time was 72 h.(6)The indirect ELISA methodshowed that both the inactivated vaccine group and the empty capsid immunized group produced lower levels of FMDV neutralizing antibody and High levels of PPV specific antibodies.(7)The indirect ELISA results were analyzed by SPSS software,and the immunogenicity of PPV empty capsid was further verified.