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Objective To prepare the monoclonal antibody (mAb) against the African horsefever virus (AHSV) VP7 protein and to identify it. Methods mAbs were prepared by using baculovirus expressed VP7 protein in BALB/c mice, and the effect of mAb was detected by ELISA, indirect immunofluorescence assay (IFA), and AHSV positive serum blockade. Results Four mAb strains were selected, including 20A8, 28B3, 30G8 and 47E6, among which 47E6 had the best blocking effect. Conclusion mAbs were successfully prepared against VP7 protein.

Objective: The present study was conducted to prepare and characterize a monoclonal antibody (mAb) against the nucleocapsid (N) protein of Schmallenberg virus (SBV)., Methods: The SBV N gene was cloned into pET-28a-c(+ and pMAL-c5X vectors and then transformed into E.coli BL21. Histidine (His)-tagged (His-SBV-N) and maltose-binding protein (MBP)-tagged (MBP-SBV-N) fusion proteins were respectively induced to express by IPTG and purified by nickel-nitrilotriacetic acid (Ni-NTA) agarose and amylose resin. His-SBV-N was used to immunize BALB/c mice to prepare mAb, and MBP-SBV-N was used as the coating antigen in ELISA to screen mAb-secreting hybridomas and to determine mAb titers. The mAb against SBV N protein was purified from the ascitic fluids using protein G sepharose. Western blotting and indirect immunofluorescence assay were utilized to analyze the reactivity and specificity of the mAb., Results: One mAb specific for SBV N protein (named 1F2) was successfully screened and purified. The titer of 1F2 was 1:32 000. Besides, the isotype of 1F2 was determined to be IgG2α/κ. 1F2 reacted with both recombinant SBV N proteins and SBV isolates. It was also cross-reactive with the N proteins of genetically related Shamonda, Douglas and Akabane viruses, but not with the Rift Valley fever virus N protein., Conclusion: One mAb specific for the SBV N protein was successfully prepared, it provides a useful tool for the serological detection of SBV.

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.