Ulbp 4

For surface marker staining of γδ T cells, the following anti-human antibodies from BioLegend (unless otherwise indicated) were employed: TCRγδ PE (clone B1, 1:25); TCRγδ PE (Miltenyi, clone REA591, 1:10); TCRγδ BV421(clone B1, 1:10); TCR Vδ1 FITC (Miltenyi, clone REA173, 1:10); TCR Vδ2 PE (Miltenyi, clone 123R3, 1:100); TCR Vδ2 PerCP (clone B6, 1:25); CD27 AF700 (clone M-T271, 1:25); CD27 APC (clone M-T271, 1:25); CD45RA FITC (clone HI100, 1:25); CD69 AF700 (clone FN50, 1:4); CTLA-4 APC (clone L3D10, 5 μl); and PD-1 BV421 (clone EH12.2H7, 1:20).
For breast cancer cell line surface staining, anti-human MICA/B PE (clone 6D4, 0.1 μg); ULBP-2,5,6 (R&D systems, clone 165,903, 0.2 μg); ULBP-3 (R&D systems, clone 166,510, 0.04 μg); ULBP-4 (R&D systems, clone 709,116, 0.1 μg).

The following anti-human antibodies for staining of cell surface markers were used: CD3 (HIT3a), CD33 (WM53), CD45 (HI30), CD34 (561), CD112 (TX31), CD155 (SKII.4), MIC-A/B (6D4), Annexin V, and 7AAD, which were all purchased from BioLegend, and ULBP4 (709116) from R & D Systems. Data acquisition was performed using C6 Accuri (BD Biosciences), LSRII (BD Biosciences), or Attune NxT (ThermoFisher) flow cytometers and data were analyzed using FlowJo version 10.