Sytox dna binding dye

CLBL‐1 and MDCK cells were treated with AZD1480 and CYT387 as described in the cell viability assay. JAK inhibitor‐treated CLBL‐1 and MDCK cells were stained with Annexin V and SYTOX Red dead cell staining and were evaluated by flow cytometry. After 72 hours of drug treatment, cells treated with DMSO, AZD1480 (2 μm and 5 μm), or CYT387 (1 μm and 5 μm) were collected and washed once with Dulbecco's phosphate‐buffered saline (DPBS) and once with 1 × Annexin V binding buffer (0.1 m Hepes/NaOH, 1.4 m NaCl, and 5 mm CaCl2). Cells at a concentration of 1 × 10⁶ cells/mL were resuspended in the binding buffer solution and stained with fluorescein isothiocyanate (FITC)‐conjugated Annexin V (eBioscience19; 1 : 40) and SYTOX DNA binding dye (Thermo Fisher Scientific10; 1 : 1000). Samples were analyzed with BD LSRII flow cytometry machine (BD Biosciences17), and results were analyzed by FlowJo v10.0.7 software18.