Preparation of the anti-AIV(H9)-idiotypic monoclonal antibody and the investigation on its immunogenicity

The SPF chickens were inoculated with avian influenza virus, subtype H9N2 to prepare the high titer serum antibodies with the subsequent purification with precipitation by saturated (NH4)2SO4. Thus prepared IgG emulsified with Freund's adjuvant was used as antigen to immunize SPF BALB/c mice, and the spleen cells of the immunized mice were fused with myeloma cells NS-1 by PEG to prepare the hybridoma cells. By using this procedure, three strains of hybridoma cells secreting the specific monoclonal antibodies could be detected by indirect ELISA assay, and they were designed as 2H1D1, 2H1D4, 3H2D7 respectively. The antibody titer of the monoclonal antibody in the supernatant in the cell cultures of these 3 strains of hybridoma cells was 2-4, and that from the ascetic fluid was 300～500 times than that of the supernatant of the cell cultures as demonstrated by indirect ELISA. It was also found that the monoclonal antibodies thus prepared could react specifically with chicken anti-AIV subtype H9N2 serum. Two high titered anti-idiotypic antibodies secreted by the 2H1D1 showed excellent neutralization reaction upon AIV on MDCK cells, and the PD50 were 10-1.8/10 μl and 10-1.6/10 μl respectively. It demonstrates that the anti-idiotypic antibody vaccine shows superior immunogenicity and can induce more neutral antibodies in SPF chickens than the inactive vaccine.