Preparation And HRP-labelling Of 19 Rabbit Antibodies Against Avian IgYs

Animals are a reservior of pathogens of infectious diseases and emerging infectious diseases. The dedective methods to these animals carring pathogens or suffering from the disease usually are PCR, DNA array and serological detecting technique. PCR and DNA array are very sensitive, but it can only detect the tissues being infected. The serological detecting technique based on antibodies could be used more widely to the animals being infected and after infection, although, this technique showed a low specifity compared with PCR and DNA array. Therefore, the serological detecting technique is a very important method for the monitoring of infectious diseases and emerging infectious diseases. The problem is that the resource of secondary antibodies is so limited in the world and the resource is needed urgently for the monitoring of infectious diseases and emerging infectious diseases over the world.The purpose of the current paper was to collect and purify the IgY of birds closing to human and produce the HRP labeled sendary antibodies against birds' IgY to set up the resource for the monitoring of infectious diseases and emerging infectious diseases.In the current paper, 19 kinds of bird eggs were collected and purified with different methods as water diluted method, affinity chromatograph, PEG precipitation, ion exchange chromatography, class ammonium sulfate precipitation. The purified IgYs were subjected to the rabbits to produce the antisera which were identified by double immunodiffusion and western blotting. The secondary antibodies were then purified by Protein-A affinity chromatography from the antisera and conjugated with horseradish peroxidase using the improved method of NaIO₄. The working condition of the labeled antibodis was determined by direct ELISA. The specificity of the labeled antibodies was tested by western blot.So far, the IgYs from 19 spieces of birds were purified and the secondary antibodies were prepared, which include chicken, goose, little grebe, purple heron, night-heron, anser, common shelduck,red-crested pochard, quail, peacock, black-wingedstilt, sparroew, ostrich, cormorant, tern, pigeon, parrots, ladyhood-chicken. Senventeen of them were labled with horseradish peroxidase.