Cd56 b159 pe cy7

All antibodies used in this study were purchased from BD Biosciences unless stated otherwise. The clone numbers are indicated in parentheses. PBMC were harvested for flow cytometry after 24 and 48 h post-establishment of the co-culture. Cells were first washed with PBS and incubated at room temperature for 15 min with Fixable Live/Dead Blue Viable Dye (Thermo Scientific). After incubation, the cells were washed in PBS buffer containing 0.5% BSA and 2 mM EDTA. Subsequently, the cells were incubated with the following antibodies: Vδ1 TCR (TS-1, Thermo Scientific) FITC, Vδ2 TCR (B6, Biolegend) PerCP, CD3 (UCHT1) V500, CD8 (SK1) APC-Cy7, CD14 (MϕP9) PE-CF594, CD56 (B159) PE-Cy7, CD69 (FN50, Biolegend) BV421, CD83 (HB15e) APC, CD161 (HP-3g10, Biolegend) BV605, Vα7.2 (3C10, Biolegend) PE, and CD38 (HB7) BUV395 for 15 min at 4°C. Assays on NK cells were performed additionally with the following antibodies: CD3 (UCHT1) V500, CD16 (3G8, Biolegend) FITC, CD56 (B159) PE-Cy7, CD38 (HB7) BUV395, CD69 (FN50 Biolegend) PE. Flow cytometry analyses were performed using a BD LSRII flow cytometer (BD). Subsequent gating analyses were performed using FlowJo software version 9.9.6 or 10.5.2 (Tree Star Inc.).