Expression of ADRB2 in human PBMCs, enrichement of pDCs.

<p>(A, B) PBMCs were generated from freshly-drawn blood from healthy human donors. After staining with antibodies against CD123, CD304, CD14 and ADRB2 or an isotype control, cells were incubated with FITC-labeled secondary anti-rabbit-IgG antibody, and ADRB2 expression was assessed by flow cytometry. (A) Histograms showing the fluorescence signal for ADRB2-FITC in PBMCs and PBMC subsets. B) Percentage of ADRB2⁺ cells within pDC- or monocyte-subpopulations within PBMC. (<b>C</b>, <b>D</b>) PBMCs were isolated from freshly prepared buffy coats from healthy human donors and pDCs were subsequently enriched by MACS technique. (<b>C</b>) Cells were labeled with anti-human-CD303 antibody, and the frequency of CD303⁺ cells was determined by flow cytometry. Dot plots show percentage of CD303⁺ cells before (upper panel) and after MACS procedure (lower panel) (gated on live cells). Right: Histogram showing the fluorescence signal for CD303 before and after MACS procedure. (<b>D</b>) Following MACS procedure, enriched pDCs and the pDC-depleted PBMCs were stimulated with PBS (vehicle) or CpG ODN 2336 (2.5 or 5.0 µg/ml). After 24 hours, IFNA1 release into the supernatant was measured by ELISA; p<0.005 for enriched pDCs vs. pDC-depleted PBMCs. CpG = CpG ODN 2336; SSC = side scatter. *** p<0.005.