TLR signaling ex vivo. (A) ISG expression was measured in controls and patients, using either a 27 (left) or 6 (right) ISG panel (measured on a NanoString platform or using RT-qPCR, respectively) to calculate an IFN score. Colors denote individuals. Values correlate to data presented in Table S1. (B–E) Ex vivo stimulation of blood cells. (B and D) IFNα and TNF production following stimulation of TLR7 (CL264 5 µg/ml), TLR8 (TL8-506 10 ng/ml), and TLR7/8 (R848 0.5 µg/ml), in bulk PBMCs. (C and E) IFNα and TNF production following stimulation of TLR7 (CL264 5 µg/ml), TLR8 (TL8-506 10 ng/ml), and TLR7/8 (R848 0.5 µg/ml) in sorted monocytes. Cells were extracted from healthy individuals (controls), one symptomatic patient (G325C-P2) and one asymptomatic individual (G325C-P1) heterozygous for the G325C substitution (B and C), and two clinically symptomatic patients (R525P-P1 and R525P-P2) heterozygous for the R525P substitution (D and E) in UNC93B1. Mean ± SEM of two to three experiments with individual patient data represented by symbols and pooled according to mutation. Two-way ANOVA with Sidak’s post-hoc test (NS, CL264, and TL8-506) or Mann–Whitney test (R848): ****P < 0.0001, **P < 0.01, *P < 0.05. Source data are available for this figure: SourceData F3.