Purification And Application Of Immunoglobulin Against CDV, CAV, CPV For The Special Economic Animals

The purposes of this study is to find good methods for getting a quantity high titer serum and immunoglobulin, decrease the produce cost and control the infectious diseases. The study was compose of three sections:Test l:The selection of immune programs and the extraction of immunoglobulin against canine distemper, canine parvovirus, canine adenovirus. The test was studied in two foxes field. 9 programs were designed for three diseases. The serum samples of foxes were gathered after the last vaccinated. The liters of the neutralization antibody against CDV and hemagglutination inhibitation antibody against CPV, CAV were detected. The result manifest that there was no significant between over twice reinforcement vaccination and once reinforcement vaccination, on the base of background titers were no significant between the two field. The same result was observed in CPV and CAV groups. When three vaccines were used respectively, their serum titers were higher than those when they were combined. But the whole level of serum titer in combined vaccine group was higher than others.IgG was extracted by salting out with ammonium sulfate and purified by ion exchange chromatography with DEAE cellulose. The purity was detected by SDS-PAGE, and only IgG's heavy chain and light chain protein tope were showed up. The activity of IgG was tested by technique of immune credeschs gold. The gold particles and virus could be seen binded together in the electronic microscope, which indicated the activity of purified IgG was high.Clinical application of hyperimmunalserun was used to treat dogs with clinicalsigns compatible with canine distemper and parvovitus enteritis. The result showed that serum against CDV was utility to dogs in the CD initial stage, and the survival rate for CP dogs was 87.5% after treated by hyperimmunalserum.Test 2:Extraction of egg yolk immunoglobulin against CD, CAV. 20 hens were allotted to two groups at randomly , 10 hens per group. One group was immunized with CDV vaccine, and the other group was CAV. Serum samples and eggs were collected weekly for detected antibody titers. High titer egg yolk was collected for following IgY extraction. IgY was extracted from diluted egg yolk with different fold water, and 10-fold dilution with pHS.O water was manifested the best way that lipids can be eliminated from the supernatant. Then 7 different methods for extracting IgY were compared. The result of SDS-PAGE certificated the method 6%octanoic acid combined with ammonium sulfate got the highest purity, the next was alginic acid sodium combined ammonium sulfate. Except the IgY got from by the method 6%octanoic acid combined with ammonium sulfate, there were over two tape in others SDS-PAGE electrophoretogram. Different molecular weight IgY exitence in the purified IgY suggested that the IgY is uniform.Through colligate comparison, the best method suit for industrialization is eliminating lipid by 0.07% alginic acid sodium combined with ammonium sulfate salting out afterl0-fold diluted with pH5.0 water.Animal safety test manifested that the egg yolk from vaccinated with CDV and CPV hens was safe to mice and rabbits.Test 3:Detected activity of serum and immunoglobulin samples by Indirect ELISA test, rabbit antibody against foxes'IgG and hen's labeled with HRP IgY was used in indirect ELISA test. The result of ELISA displayed that the all the serum against CDV and CAV, and immunoglobulin against CAV had high activity. But the IgG against CDV didn't appear a high activity.