Flow cytometric analysis reveals that leukemic pDCs express CD33 at different levels. Leukemic pDCs from 8 other patients (patients no. 2-9, noted as P2, P3, P4, P5, P6, P7, P8, and P9, respectively) were stained with either anti-CD33 mAb (hatched gray curve) or corresponding isotype control mAb (open curve) and analyzed by flow cytometry. Four different anti-CD33 mAb clones were used: 2 were conjugated with PE (LeuM9 or WM53) and 2 with FITC (WM-54 or D3HL60.251). On each histogram, percentage of CD33⁺ cells (defined using isotype control mAb staining) and relative fluorescent intensity (rfi; corresponding to MFI R = MFI obtained with anti-CD33 mAb/MFI obtained with isotype control mAb) are indicated. According to Garand and Robillard,²⁹  leukemic cells are considered positive when the percentage of positive cells is more than 20% or rfi is more than 2. Due to limited biologic material, cells from patient no. 9 were stained only with PE-conjugated anti-CD33 mAb. Cells from patient no. 1 (not shown) presented the same pattern of staining as cells from patient no. 3 (ie, 30% of positive cells with 4.3 rfi after PE-conjugated LeuM9 mAb and 8% of positive cells with 1.5 rfi after FITC-conjugated D3HL60.251 mAb). Cells from 2 patients presenting CD33^- ALL (ALL no. 1 and ALL no. 2) stained with PE-conjugated anti-CD33 mAb (LeuM9 and WM53) were also shown as control (Ctrl). In these cases, rfi was always less than 1. Purified cells from patient no. 2 expressed CD33 at the highest levels and were previously shown to produce IFN-α in response to inactivated influenza virus (1302 and 1695 IU/mL in 2 independent experiments after 2 separate thawing procedures).¹⁷  Purified cells from patients no. 4, 7, and 8 also produced IFN-α in response to virus (56, 65, and 953 IU/mL, respectively, versus < 0.6 IU/mL in unstimulated conditions).¹⁸