Gating strategy for the detection of the investigated Tregs. PBMCs were extracellularly stained with PerCP-labeled anti-human CD4-, PE-labeled anti-CD25 antibody, Brilliant Violet (BV570)-labeled anti-CD127 antibody and intracellularly stained with Alexa Fluor (AF488)-labeled anti-human FoxP3 antibody. Lymphocyte population was gated according to forward scatter (FSC) characteristics and side scatter (SSC) characteristics (A + D). Gated lymphocytes were then separated in CD4 + CD25high cells (B + E) and CD4 + CD25highCD127lowFoxP3+ cells (C + F, named Treg). Representative results of a healthy control (A-C) and a patient with neuropathic pain (D-F) are shown. Regulatory T-cells, Tregs; PBMC, Peripheral blood mononuclear cells; Forkhead-Box-Protein 3, FoxP3.