Preparation Of AIV Inactivate Vaccine Adsorbed By Nano-particulated Aluminum Hydroxide Adjuvant

Avian influenza is a acute and strong infection disease caused by influenza virus type A adapted to birds.The nation force-proposal defined,once the HPAI outbreaked,all of the poultry will be killed in 3 km surrounding of the epidemic focus,vaccine in an urgent manner in 5 km surrounding of the epidemic focus,so that building a isolation zone of immune.At present,the routine way to prevent and control HPAI is to inject oil emulsion vaccine,which,however,turns out unsatisfactory. But it takes quite long to reach effective antibody titer after the first vaccine,which makes it difficult to achieve immune protection within a short time in emergency.Oil adjuvant,with strong stimulation,is difficult to absorb and some is even remained in the body when the poultry are killed, which will great effect on the poultry products quality.United States Federal Government regulated that cannot inject the emulsion vaccines in 42 days before sold.Therefore,it's in urgent need to search an high effectiveness,safe and new adjuvant which is easy to vaccine in emergency.In this study,we prepared the nano-particulated aluminum hydroxide adjuvant,and on this basis to explore the condition of AIV inactivate adsorbed by nano-particulated aluminum hydroxide adjuvant,which is the best concentration,and the quality of the vaccine.The main contents are as follows in this subject:1 Preparation and identification of nano-particulated aluminum hydroxide adjuvant(NAHA)The particle of NAHA was prepared Bybenzalkonium Bromide-Glycerin-Cyclohexane- AlCl3 W/O type microemulsion.The mean NAHA diameter is 197.2nm identified by Zetasizer 3000HS and the content is 2mg/mL by titration technique.Accordingly it provide a material basis to further check the immune effect of nanoparticulate alum adjuvant.2 Preparation of AIV inactivate Vaccine adsorbed by nano-particulated aluminum hydroxide adjuvantWe chose HA,fluorescence RT-PCR,SINGLE-RADIAL-DIFFUSION to ASSAY antigen in allantoic fluid.HA=9Log2,the result of fluorescence RT-PCR is mast,content of hemagglutinin is hemagglutinin 3.133μg/ml.The three methods may evaluate absorption rate effectually.In the best absorbing surroundings,we prepared the high absorption rate AIV vaccine.HA,fluorescence RT-PCR,titration technique determined the sedimentation rotation speed of antigen and adjuvant respectively.To allantoic fluid and adjuvant volume ratio of 1:2 mixed with the selection of different temperature,adsorption time and adsorption adjuvant concentration.And centrifugal,supernatant assayed by HA,fluorescence RT-PCR and complexometric titration method.The result is that 7000rpm is centrifugal speed to detect absorption rate.When the aluminum hydroxide final concentration is 0.700mg/mL in vaccine,adsorption in 2h 25℃,antigen adsorbed by nano-particulated aluminum hydroxide adjuvant completely,while conventional one incomplete.3 Detected quality of the vaccineIn view of product development requirements,we detected the above-mentioned processes of avian influenza inactivated vaccine,the results show to meet the national standard,which is propitious to stored in 4℃low temperature environment and nano-aluminum hydroxide adjuvant vaccine is more stable than conventional one.Conclusion:This study confirmed the nano-aluminum hydroxide is a good immune adjuvant, can effectively adsorbed avian influenza virus.Reported best adsorption conditions and the adjuvant concentration of nano-aluminum hydroxide adsorption AIV H5N1 for the first time,and the adsorption rate of nano-aluminum hydroxide is higher than conventional group.Quality of nano-aluminum hydroxide vaccine can be controled,to provide a solid theoretical foundation for the improvement and development of adjuvant,to provide a theoretical basis for the mechanism of adjuvant,to lay a solid foundation for further clinical trials.