Preparation of a new lot of anti-classical swine fever virus monoclonal antibody for distribution by the national veterinary assay laboratory
2007
Aoki, H.(Japan. Ministry of Agriculture, Forestry and Fisheries, Kokubunji, Tokyo. National Veterinary Assay Lab.) | Sekiguchi, H. | Kozasa, T. | Nakamura, S.
A new lot of anti-classical swine fever (CSF) virus monoclonal antibody 3D11 (mAb) for use in the quality control assay (virus content test) for CSF live vaccines was prepared and the concentration of the mAb was determined. Hybridoma 3D11 cells that grew well and showed high-1evel mAb production were isolated after the cells had been grown and recloned twice. The mAb secreted by these cells was confirmed to react with PK15 cells infected with the CSF vaccine strain, GPEsup(-), by the indirect immunoperoxidase (IIP) method. To determine an appropriate dilution for the quality assay, pooled ascites were exposed to PK15 cells infected with l0E2 TCIDsub(50) of the vaccine strain and evaluated by the IIP method using transitions of absorbance (dual wave length: 492/630 nm). Ascites diluted more than 3,000-4,000 times showed a clear positive reaction without non-specific binding. Moreover, the virus titer obtained by the IIP method using the ascites diluted less than 3,000 times corresponded to titers observed using both the interference and the direct-CPE methods. After IgG was purified from the ascites, absorbences obtained by the IIP method were compared between the ascites and purified IgG. The purified IgG showed the same absorbancy as the ascites. Our results suggest that the new lot of ascites containing anti-CSFV mAb can be used at a 3,000-fold dilution for the quality control assay (virus content test) for CSF live vaccines.
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