Preliminary Study Of Adjuvant Th Of Foot-and-Mouth Disease Vaccine

Foot-and-mouth disease (FMD) is a highly infectious viral disease which is caused by foot-and-mouth disease virus (FMDV). It is characterized by fever and vesicles in the mouth and on the muzzle, teats, and feet. FMDV is a single stranded positive sense RNA virus belonging to the genus Aphthovirus in the family Picornaviridae. FMDV contains one ORF which encodes four structural proteins VP1, VP2, VP3and VP4, and seven nonstructural proteins2A,2B,2C,3A,3B,3C and3D.Nonstructural protein3D is a RNA polymerase, involving in RNA replication. Multiple Th epitopes capable of helping B cell activation have been identified on3D protein. These epitopes facilitate the development of novel FMDV vaccines and adjuvants.In this study, we aimed to obtain a recombinant protein harboring multiple Th epitopes from3D protein and to examine its effect on the activation of the immune system. The genes encoding these epitopes have been synthetized and cloned into the prokaryotic expression vector pET28a and vector pAIYM. Two positive recombinant plasmids pET28a-Th115and pAIYM-Thl57were constructed and transformed into E. coli BL21. Upon induction with IPTG, recombinant proteins Th115(16KD) and Th157(19KD) were successfully expressed respectively. Western blot indicated that Th115and Th157could react specifically with His monoclonal antibody. After treatment of sonication, the supernatants of Th115and Thl57were purified through Ni-NTA chromatography. Purified Th115and Th157were obtained with high concentrations.After that, Purified Th115and Th157protein were employed as adjuvants for immunization with vaccine against FMDV in clinical trials. We designed for experiment groups:vaccine-Th115adjuvant, vaccine-Th157adjuvant, vaccine alone, and PBS groups. ELISA was used to measure antibody response. The results showed that the antibody titer of vaccine alone group was1:12800, the antibody titer of vaccine-Th157adjuvant group was1:51200and the antibody titer of vaccine-Th115adjuvant group was1:102400. MTT method was used to deteced T cell proliferation, the results show that more T cell proliferation was found in vaccine-Thl15adjuvant and vaccine-Th157adjuvant-groups than the group of vaccine alone. Thus, Th115and Th157have the capacity to enhance the activation of the immune system, and have the potential to be used as novel adjuvants in the preparation of FMDV vaccines.