The Screening Of Adjuvant Applicable For Pig-original ETEC Pily Vaccine For Laying Hens

For screening immunologic adjuvants of high efficiency,security,low cost,easyinjection,low side effect suitable for egg-laying hen,this experiment will take ETECpilin(K₈₈,K₉₉and⁹⁸⁷P) as model antigen,respectively blended withSPA,SPB,SPC,ISCOMs,Freund's Adjuvant,Astragalus mongholicus polysaccharideand propolis to make pig-original ETEC multivalence pily vaccine,immuned browncrust egg-laying hen(From American Hailan),analyzing the egg-layingfunction,overall outiside apperance and the influence for tissue of injectingpart,discussing the otherness of regular pattern of which conduct the growth anddecline of serum antibody and yolk antibody.The main researching consequeceabstract following below:1. Make multivalence pili vaccine of each group:Adopt60～65℃water verbationto extract K₈₈,K₉₉and⁹⁸⁷P,by purification,inspection and analysis of SDS-PAGEelectrophoresis,the consequece indicating that successfully making K₈₈,K₉₉ and⁹⁸⁷P three kinds of pili protein.Blending three kinds of pili protein as specificproportion and take the compound as model antigen,then blend the compound withseven kinds of immunoadjuvant to make vaccine as the respective designatedprocedure,do inspection for made vaccine in these aspects ofasepsis,appearance,color,stablization(whether layered or not),etc.The consequenceindicating: Inoculating made vaccine into the ordinary nutrient broth,aftercultivation,there is no layer caused);ISCOMs adjuvant vaccine presenting whitenesslike water,transparency,no layer; Astragalus mongholicus polysaccharide injected intoISCOMs adjuvant vaccine presents the tawny which is transparent,no layercaused;propolis adjuvant presents puce,semitransparency and deposit.,the vaccinemade could be applied to immune of egg-laying hen.2. ETEC subunit vaccine processed by different adjuvant are injected into egg andthe effect to performance of laying egg:After making the immune of the ETECmultivalence pili vaccine egg-laying hen injected withSPA,SPB,SPC,ISCOMs,freund's adjuvant, Astragalus mongholicus polysaccharide and propolis adjuvant,the laying rate is lower in1～3days after immune to eachgroup of egg-laying hens,after a while,the rate would restore;Apart from SPBgroup,the averange of laying rate is between71.37%and83.15%; in othergroups,higher than70%of pre-immunation,the laying rate of SPB group is average50.25%,evidently lower than the layring rate of other groups(p<0.05).The conferenceindicate that SPB adjuvant could decreasing the function of laying by comparisonwith other adjuvants.3. The consequece of serum anti-body of each group inoculated subunit vaccine ofETEC made by different adjuvant:After injecting three times,the valence of the serumantibody in7groups from top to low successively is:freund'sadjuvant,SPC,SPB,SPA,ISCOMs, Astragalus mongholicus polysaccharide andpropolis adjuvant;SPC and Freund's adjuvant are sustaining immune with themaximum time,secondly is SPB,SPA and ISCOMs,the minimum time is forAstragalus mongholicus polysaccharide and propolis adjuvant.4. The consequece of yolk anti-body of each group inoculated subunit vaccine ofETEC made by different adjuvant:After injecting three times,the valence of the yolkantibody in7groups from top to low successively is:freund'sadjuvant,SPC,SPB,SPA,ISCOMs, Astragalus mongholicus polysaccharide andpropolis adjuvant;SPC and SPB are sustaining immune with the maximumtime,secondly is Freund's adjuvant,SPA and ISCOMs,the minimum time is forAstragalus mongholicus polysaccharide and propolis adjuvant.5. The situation of entire appearance of egg-laying hens in each group and thepartial anatomy to injecting part inoculated with ETEC subunit vaccine made bydifferent adjuvant:The observed result indicates after injecting vaccine,obviousadverse reaction don't appear in the hens of other groups apart from the hens injectedin SPB and SPC adjuvant sustaning2h heating reaction;All the hens are better indrinking,eating,appearance and mental situation;Randomly pick up5hens from eachgroup after3times immune,dissect the part injected,could see the clear inflammatoryinfiltration,axunge nodule abscess in the injected part with SPB,SPC and Freund's adjuvant,the ratio is2/5,5/5,4/5respectively,there are no side effects in othergroups(including control group).To sum up,we could get the conclution below:by comparison with other6groups,the vaccine combined with SPA adjuvant and module antibody immunizinghens could efficiently improve the immune valence of serum antibody and yolkantibody and sustain the high titer above30days since immune ends;and the adjuvantvaccine makes little impact on the property of laying;After injecting the adjuvantvaccine,there is no obvious side effects,great security;there is no obvious impact onappearance of hens and muscular tissue injected,couldn't influence the value ofeating;the adjuvant is easy to be made and injected with lower cost and easyoperation,which is a kind of immune adjuvant suitable for egg-laying hens ofhigh-efficiency,safe,low cost,easy injection and little side effects.The consequence ishelpful to opt for fitted adjuvant and make multivalence pili vaccine to get a largeamount of yolk antibody with high purity,used to prevent the diarrhea caused by pigletETEC.