Pe conjugated anti cd158b

K562 cells (China Center for Type Culture Collection, Wuhan, China) were used as target cells in the NK cytotoxicity assay. K562 cells were cultured in RPMI 1640 (Gibco, Grand Island, NY, USA) supplemented with 10% fetal bovine serum (FBS; Hyclone, Logan, UT, USA). 3,3′-Dioctadecyloxacarbocyanine perchlorate (DIO) (1911717; Invitrogen, San Diego, CA, USA) was dissolved in dimethyl sulfoxide (Sigma, St. Louis, MO, USA) to a concentration of 3 mM. Propidium iodide (PI; MKCB0899V; Sigma) was used to detect apoptosis.
Flow cytometry to identify NK cells and subpopulations expressing certain markers and effector molecules was performed using the following fluorophore-conjugated monoclonal mouse anti-human antibodies: PerCP-conjugated anti-CD3 (347344; BD Pharmingen, San Jose, CA, USA), PE-cy7-conjugated anti-CD56 (557747; BD Pharmingen), PE-conjugated anti-NKp30 (12–3379; eBioscience ThermoFisher, Waltham, MA, USA), APC-conjugated anti-NKp46 (17–3359; BD Pharmingen), FITC-conjugated anti-NKG2D (11–5878; eBioscience), FITC-conjugated anti-CD158a (556062; BD Pharmingen), PE-conjugated anti-CD158b (559785; BD Pharmingen), PE-conjugated anti-granzyme B (561142; BD Pharmingen), Alexa Fluor 647-conjugated anti-perforin (563576; BD Pharmingen), and Alexa Fluor 488-conjugated anti-granulysin (558254; BD Pharmingen).