Development and evaluation of a competitive enzyme-linked immunosorbent assay using a monoclonal antibody for diagnosis of severe fever with thrombocytopenia syndrome virus in bovine sera

Lee, Hyojin; Kim, Eun-Ju; Song, Jae-Young; Choi, Jeong Soo; Lee, Ji Youn; Cho, In-Soo; Shin, Yeun-Kyung

The gene for the recombinant NP was amplified from the commercially synthesized S fragment of SFTSV (GenBank accession No. HQ141612) and cloned into a protein expression vector. A 34 kDa recombinant protein band was well maintained after all purification procedures (Fig. 1 ). This recombinant NP was used as the antigen to generate polyclonal and mAbs. NP-specific polyclonal antibody was obtained in rabbits after four antigen immunizations, and the titer of the antibody was tested by IFA and Western blotting (Fig. 2) . NP-specific mAbs were produced from the spleen hybridoma cells of immunized mice, and the three clones (6D55, 8F31, and 10G7) showed strong interactions in the IFA and Western blot tests; therefore, these clones were used as competitive antibodies to develop the cELISA. A clear band corresponding to the size of NP, which appeared only after SFTSV infection, was observed upon Western blot analysis with all generated antibodies (panel A in Fig. 2) . Moreover, IFA using these antibodies showed intense cytoplasmic staining, specifically in Fig. 2) .