Annexin 5 fitc pi double staining kit

1 × 10⁶ transfected cells (NCI-H1299 and NCI-H1650) were prepared and seeded in 6-well plates for cell apoptosis assay, in presence of Annexin V-FITC/PI double staining kit (Invitrogen). After 15 min of double-staining in the dark, cells were subjected to flow cytometer, as instructed by provider (BD Biosciences, Franklin Lakes, NJ, USA). Bio-repeats were run in triplicate.

An annexin V-FITC/PI double-staining kit (Invitrogen, Life Technologies GmbH, Darmstadt, Germany) was used for apoptosis detection. In brief, CCRF-CEM cells were seeded in 6 well plates at a density of 1 × 10⁶ cells/well. They were then treated with increasing concentrations of the artemisinin dimer isoniazide (2.5, 5, and 10 µM) or DMSO as a negative control and incubated for 48 h. Afterward, the cells were harvested, washed, and resuspended in 1 mL cold PBS combined with 500 µL binding buffer. Subsequently, the cells were stained using 5 µL annexin V-FITC for 15 min, followed by the addition of 10 µL of propidium iodide (PI) for 15 min at room temperature in the dark. Finally, apoptosis was detected using a flow cytometer (BD Accuri™ C6, BD Biosciences, Becton Drive, Franklin Lakes, NJ, USA). The signal detector for FITC is FL1, while for PI it is FL2. This experiment was repeated three times.