Eleven Ifnar1-deficient 129S2/SvPas mice were either infected i.p. with 10 4 PFU rZHDNSs-GFP RVFV (N = 6) or mock-infected (N = 5). Twenty-four hours later, the spleen was harvested. Erythrocytes were lysed using NH 4 Cl (9 g/L) buffer. The rat anti-mouse CD16/CD32, clone 2.4G2 antibody (BD Pharmingen, San José, CA) was used to block non-antigen-specific binding of immunoglobulins to Fc-receptors. Cells were stained using a combination of the following antibodies: (i) PE-conjugated rat anti-mouse NKp46/CD335 (BD Pharmingen). (ii) PerCP-Cy5.5-conjugated hamster antimouse CD3 (BD Pharmingen). (iii) APC-conjugated rat anti-CD19 (BD Pharmingen). (iv) Pacific Blue-conjugated rat antimouse CD11b/Mac-1 (eBioscience, San Diego, CA). (v) APCconjugated hamster anti-mouse CD11c/Itgax (BD Pharmingen). (vi) Alexa Fluor 700-conjugated rat anti-mouse MHC Class II (I-A/I-E) (eBioscience). (vii) PE-conjugated rat antimouse Ly6G/Gr-1 (BD Pharmingen). (viii) Biotin-conjugated anti-mouse CD115/c-Fms (eBioscience) with Streptavidin-PerCP-Cy5.5 (BD Pharmingen) as second-step reagent. All staining procedures were conducted on ice. Then, the cells were fixed with 4% formaldehyde. Fluorescence was measured using a FACSAria II flow cytometer (BD Biosciences, San Jose, CA), and data analysis was performed using CellQuest (BD Biosciences) and FlowJo (Ashland, OR) softwares. Dead cells were visualized using the Fixable Aqua Dead Cell Stain kit (Invitrogen, Carlsbad, CA). Fluorescence compensation settings for multicolor flow cytometric analysis were optimized based on single-stained polystyrene microparticles (Comp-Beads, BD Pharmingen).