Cd3 brilliant violet 785

Cells were phenotypically analysed using the Beckman Flow Cytometer. To exclude dead cells from the analysis, 7-amino-actinomycin-D was added to the cells prior to acquisition. For 9-color analyses on the BACKMAN, the following conjugated mAbs were combined: CD56-Brilliant Violet 605 (catalog number: 362538, BioLegend, China), CD16-Brilliant Violet 510 (catalog number: 302048, BioLegend, China), CD45-APC-Cyanine 7 (catalog number: 368516, BioLegend, China), CD3-Brilliant Violet 785 (catalog number: 344842, BioLegend, China), NKG2A-FITC (catalog number: 130-113-565, BioLegend, Miltenyi, Germany), CD8-PB450 (catalog number: 562428, BD, USA), CD4-APC (catalog number: 5553492, BD, USA), CD33-PE (catalog number: 555450, BD, USA), IFN-γ-FITC (catalog number: 506504, BioLegend, China), CD107a-PE-Cyanine 7 (catalog number: 561348, BD, USA), and Granzyme B-AF647 (catalog number: 560212, BD, USA). Before 9-color analyses were performed, all conjugates were titrated and individually tested for sensitivity, resolution, and compensation of spectral overlap. Isotype controls were used to define marker settings. The combinations were balanced in fluorochrome combinations to avoid antibody interactions and steric hindrance and to detect dimly expressing populations. Then, the cells were analysed using FlowJo software 10.