Study On β-propiolactone Inactivating Foot-and-mouth Disease Virus

At present,the main prevention and control measures for foot-and-mouth disease in China are prevention-oriented,and susceptible animals are regularly vaccinated.Inactivation is the most important in the process of vaccine preparation.Whether the inactivation is complete and whether the inactivation can retain more complete antigenicity is of great significance to the effect of inactivated vaccine.Nowadays,the most commonly used inactivated agent in the preparation of foot-and-mouth disease inactivated vaccine is diethyleneimine(BEI),which has a complex operation and a long inactivation time.However,β-propiolactone(BPL)has a short inactivation cycle and is relatively convenient to operate,but it is rarely used in the inactivation of foot-and-mouth disease virus.In this experiment,in order to study whether β-propiolactone can be used to inactivate foot-and-mouth disease virus and whether the immune effect of inactivated vaccine prepared after inactivating it is qualified,four strains of O/r V-1,A/r V-2,AKT-III and OHM/02 were selected for testing.Firstly,the stability of foot-and-mouth disease virus at 37℃ was verified,and then the inactivation conditions of foot-and-mouth disease virus by BPL were determined from three aspects of temperature,time and inactivator concentration.The inactivation effect was tested by sterility test,cell security test,suckling mouse security test and 146 S test after inactivation.The VP1 gene of FMDV was detected by RT-PCR.The completely inactivated virus solution was emulsified with adjuvant to prepare foot-and-mouth disease OA labeled vaccine and foot-and-mouth disease OA bivalent vaccine.The two vaccines were compared with the BEI inactivated control group vaccine from physical properties,146 S content,total protein,endotoxin and other conventional quality control items.The three vaccines were inoculated into experimental animals for side reaction verification and serum antibody levels at 14 d,21d and 28 d after inoculation were detected by ELISA kit.The results showed that FMDV was stable at 37℃ for 2h,and 146 S was not degraded.The most suitable conditions for inactivating foot-and-mouth disease virus byβ-propiolactone were 4℃,inactivated concentration of 1:1000,and inactivated for 3h.After complete inactivation,the virus solution sterility test,cell safety test,and suckling mouse safety test were all qualified.RT-PCR detection of VP1 protein gene was not affected.The results of routine quality control items and side effects of foot-and-mouth disease OA labeled vaccine and foot-and-mouth disease OA bivalent vaccine were all qualified.At 14 d,21 d and 28 d after the first immunization,the antibody level of the foot-and-mouth disease OA labeled vaccine was slightly higher than that of the control group,and the antibody level of type O and type A reached 1 : 64 at 28 d,reaching the effective protection level.Although the antibody level of the foot-and-mouth disease OA bivalent vaccine was slightly lower,it could also reach the protection level.