2.5. Quantification of dendritic cell subsets

The peripheral blood dendritic cells were measured through a flow cytometry analysis of fresh whole blood within 12 h of sampling. Briefly, whole blood cells were incubated for 30 min in the presence of the following monoclonal antibodies (mAbs) at room temperature: cocktail lineage conjugated with fluorescein isothiocyanate (Lin 1-FITC) (Caltag, Burlingame, CA, USA), anti-HLA-DR conjugated with PE-Cy5 (Caltag, Burlingame, CA, USA), anti-CD11c⁺ conjugated with PE (Caltag, Burlingame, CA, USA) and anti-CD123 conjugated with PE (Caltag, Burlingame, CA, USA). The erythrocytes were lysed using FACS lysing solution (BD Biosciences, San Jose, CA, USA) at room temperature. The cells (50,000 events) were acquired by FACScan and analyzed using the Cell Quest software (Becton-Dickinson). The dendritic cells were identified as Lin⁻ HLA-DR⁺ cells within a monocyte gate. The CD11c and CD123 expression was determined within Lin⁻HLA-DR⁺ cells to define the myeloid DC (CD11c⁺CD123⁻) and plasmacytoid DC (CD11c⁻CD123⁺) subsets.