Xi bao yu fen zi mian yi xue za zhi = Chinese journal of cellular and molecular immunology · 2013
[Preparation and characterization of monoclonal antibodies against viral hemorrhagic septicemia virus].
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Jing Hongli, Wang Jingbo, Cao Huan, Lin Xiangmei
Аннотация
OBJECTIVE: To prepare the monoclonal antibody (mAb) against viral hemorrhagic septicemia virus (VHSV) and analyze the biological properties.
METHODS: The BALB/c mice were immunized with the VHSV which was purified by differential centrifugation. Spleen cells of the immune mice were collected and fused with Sp2/0 myeloma cells by hybridoma technology. The indirect ELISA was used to screen hyridoma cells and identify the specificity of mAb. The Western blotting was used to identify antigen site.
RESULTS: After three cycle of subcloning with limited dilution method, we obtained a cell strain that secreted mAb against VHSV and was named 4A5. The indirect ELISA showed that the titer of mAb ascites was 10⁻⁴;. It belonged to IgG3 subclass κ chain and couldn't react with other fish viruses or fish cell lines in the ELISA except VHSV. Western blotting revealed that mAb 4A5 bound to a 70 kDa of protein band of VHSV as antigen.
CONCLUSION: The obtained mAb has a high specificity, which could be used for rapid diagnosis and detection of VHSV in aquaculture.
METHODS: The BALB/c mice were immunized with the VHSV which was purified by differential centrifugation. Spleen cells of the immune mice were collected and fused with Sp2/0 myeloma cells by hybridoma technology. The indirect ELISA was used to screen hyridoma cells and identify the specificity of mAb. The Western blotting was used to identify antigen site.
RESULTS: After three cycle of subcloning with limited dilution method, we obtained a cell strain that secreted mAb against VHSV and was named 4A5. The indirect ELISA showed that the titer of mAb ascites was 10⁻⁴;. It belonged to IgG3 subclass κ chain and couldn't react with other fish viruses or fish cell lines in the ELISA except VHSV. Western blotting revealed that mAb 4A5 bound to a 70 kDa of protein band of VHSV as antigen.
CONCLUSION: The obtained mAb has a high specificity, which could be used for rapid diagnosis and detection of VHSV in aquaculture.