SFTSV Infection and Antibody Detection

The IFAs were performed as previously reported [10 (link)]. Briefly, Vero E6 cells in 150 cm² culture vessels (Corning, USA) were sensitized with 2 mL of 1 × 10³ TCID₅₀/mL SFTSV for 1 h at 37℃. The infected cells were detached with 0.25% trypsin-ethylenediaminetetraacetic acid (Gibco, USA) 6 days after infection. The floating cells were spotted onto reaction wells (Paul Marienfeld, Germany) and fixed with a methanol:acetone (1:1) solution. Serially diluted serum samples were incubated with fixed cells for 1 h at 37℃, and the fluorescence was detected using fluorescein isothiocyanate (FITC)-labeled secondary antibodies. The fluorescence was observed by using a Nikon TE-2000U fluorescence microscope (Nikon, Japan). The anti-bovine and anti-horse secondary antibodies (KPL) were diluted to 2.5 µg/mL prior to use.