Hybridoma Screening and Purification of SFTS Virus Antibodies

NIH/3T3 cells were transfected with an SFTS virus Gn/Gc expression plasmid (pC030GP) [28 (link)] by using TransIT LT1, then used for immunization of BALB/c mouse to obtain antibody-producing hybridoma. Screening of hybridoma was performed with ELISA with SFTS virus-infected and virus-uninfected cell lysates (described above) as antigens. SFTSVGnSTH and SFTSVGcSTH proteins (described above) were used for immunization of GANP^® mice to obtain antibody-producing hybridoma and screening of hybridoma was performed with ELISA with each protein as antigens (Transgenic Corp., Hyogo, Japan). To obtain purified mAbs, hybridoma were cultured in Hybridoma Serum-Free Medium (FUJIFILM Wako Pure Chemical Corp.) and HiTrap™ Protein G HP Columns (Cytiva) were used. Isotypes of mouse mAbs obtained were determined with IsoStrip™ Mouse Monoclonal Antibody Isotyping Kit (Roche, Basel, Switzerland). Commercially available mouse mAbs specific to SFTS virus Gn and Gc were purchased from Immune Technology Corp. (New York, NY, USA). Two mAbs, C3A11 and C6C1 clones, were described previously [29 (link)].