Figure 1.

11-color flow cytometry gating strategies to assess circulating immune cell functionality. Dot plots represent results obtained after a healthy donor WB stimulation. (A) After short term P/I activation, we analyzed by multi-parametric flow cytometry the ability of γδ T cells (CD3⁺TCRγ9⁺), LTCD8⁺CD45RA⁺ (CD3⁺TCRγ9^negCD8⁺CD45RA⁺), memory LTCD8⁺ (CD3⁺TCRγ9^negCD8⁺CD45RA^neg), naive CD4⁺ (CD3⁺TCRγ9^negCD4⁺CD45RA⁺) and memory CD4⁺ T cells (CD3⁺TCRγ9^negCD4⁺CD45RA^neg) to synthetize IFNγ, IL-2, TNFα, IL-21 or IL-17A. (B) The “Innate Immunity” panel allowed the simultaneous identification of NK cells (LIN^negHLA-DR^negCD56⁺), DC subsets (LIN^negHLA-DR⁺) including pDC (BDCA²⁺CD11c^neg), BDCA-3⁺ mDC (CD11c⁺BDCA-3^high), BDCA-1⁺ mDC (CD11c⁺BDCA-1⁺), monocytes (HLA-DR⁺Lin⁺CD11c⁺CD56^neg) including CD14⁺CD16^+/−monocytes and non-classical (nc-monocytes CD14^lowCD16⁺).