Study On The Extract Methods And Application Of Chicken Newcastle Disease Egg Yolk Antibody

The aim of this paper is to improve the process of egg yolk antibody(IgY) extraction and purificattion, compare the effect of IgY extracted by different methods including the poloxamer methods,PEG6000 methods, the water dilution methods, and futher defat by the Freeze-thaw method.Search the methods which were simple in production, low in cost and less impact on biological activities, was screened to provide the basis for the of massive IgY.The freeze-drying method concentrated IgY, measure product biological activities with HI expermient.IgY was purified by salt precipitation with solid ammonium sulfate method, The purity and biological activities of the product was measured with SDS-PAGE and HI expermient.Highland gray cocks whose maternal antibody attenuated to almost disapper randomly assigned into 2 groups, the experiment group injected 2ml IgY including ND-Ab, AIV-Ab H5 and AIV-Ab H9.the control group injected 2 ml Sodium Chloride. Measured chicken serum antibody titer by hemagglutination inhibition test at different time.The experimentai object of this test is IgY againest NDV, cock whose Newcastle disease antibody levels was negative randomized into 6 groups, the IgY prevention group, the self-healing group, The low dose IgY treatment group, The middle dose IgY treatment group, The high dose IgY treatment group and control group, each group was inoculated NDV except the control group, begin treatment after infection 72h, observed the effect of IgY for NDV treatment.The results are as follows:1. The results of IgY extractedThe 1.0% poloxamer group settlement fast and the precipitat solid, supernatant had a little yellow; the 3.5% PEG6000 and the 1:10 water dilution method better in defat, but the precipitat was not solid, no good for operating.The 1.0% method group IgY biological activities lower 1-2 HI antibody titers than the chloroform method, the 1:10 water dilution method lower 4-5 HI antibody titers than the chloroform method, The 3.5% PEG6000 method lower 3 HI antibody titers than the chloroform method. The 1.0% poloxamer method is the best method for extraction of IgY.2. The results of IgY purifiedSaturated ammonium sulfate method could effectively purified IgY, The Purity was no difference in the 1.0% poloxamer method; the 3.5% PEG6000 method; the 1:8, 1:10 water dilution methods. SDS-PAGE showed mainly in 66KD bands, IgY biological activities increased 2-3 HI antibody titers, and the protein content increased was obvious.3. IgY product could effectively concentrated by Freeze-drying, Freeze-drying had no impact on the antibody activity, the 5 times concentrated product increased 2 HI antibody titers, the 10 times concentrated product increased 5 HI antibody titers, so Freeze-drying can improve IgY antibody titer.4. The dynamic change of Chicken serum antibody after inject IgYThe ND-Ab titer was significantly higher than the control group in 1-8 day(P <0.01),AIV-Ab H5 titer was significantly higher than the control group in 1-11 day(P <0.01),and AIV-Ab H9 titer was significantly higher than the control group in 1-14 day(P <0.01).The serum antibody titers were raised significantly just 1d post injection. The high antibody titers can continued 5-6 days and then reduce gradually.5. The results of NDV model treatmentThe cock antibody titer was negative, on protection to the NDV, but prevention group Mortality ratio was 5%, self-healing group Mortality ratio was 70%, The low dose IgY treatment group Mortality ratio was 40%, The middle dose IgY treatment group Mortality ratio was 30%, The high dose IgY treatment group Mortality ratio was 15% ,no deaths in the control group,and The IgY treatment groups better in recover, so injected IgY for NDV treatment was effective.Conclusion:1. The joint application the 1.0% poloxamer method and Freeze-thaw can be used forIgY mass production. IgY extracted by The 1.0% poloxamer method biological activities was highest, Freeze-thaw method can be used for IgY product further defat. Joint application of the two methods can be used for IgY mass production.2. Saturated ammonium sulfate method can effectively purify IgY.3. Freeze-drying method is suitable for concentrate IgY.4. Anti-NDV IgY showed significat protection effect in chicken against NDV.Inject IgY can significantly improve chicken serum antibody titer,and in IgY Passive protection againest NDV can effectively reduce mortality,the protection rate of trement groups improved 30%-55% than the self-healing group.so serum inject IgY for NDV trment is effectively.