Anti cd107a pe clone h4a3

Cells were harvested from culture washed with R0 (as for R10 but with no FBS) and rested overnight in R5 media (as for R10 but with 5% FBS). Resting ensured minimal background activation of the T-cells. On the day of activation assay, cells were harvested, and 2–5 × 10⁴ cells were incubated with 30 µM TAPI-0 (Sigma-Aldrich) (54 (link)) anti-TNF-PE-Vio770™ (clone cA2, Miltenyi Biotech) and anti-CD107a-PE (clone H4A3, BD Biosciences) Abs in well(s) of a 96U-well plate. For peptide presentation, autologous LCLs were used for clonal T-cells, and T-cell to T-cell presentation used for T-cell lines. The cells were incubated for 4–5 h at 37°C then stained with Vivid and Abs for CD3 and/or CD8, as above.

In vitro cytotoxicity experiments were performed using K562 or Beas-2B cells as the target and NK cells as effector cells. NK cells were added to K562 or Beas-2B cells with a 5:1 effector:target ratio [23 (link)]. NK cell degranulation was evaluated by CD107a staining (anti-CD107a-PE; clone H4A3; BD Biosciences) after 3 h of treatment with Golgi Stop solution (BD). Labeled cells were analyzed with a FACSCantoII flow cytometer (BD, Milano, Italy) and FlowJo software (Tree Star Inc., Ashland, OR, USA).