Annexin 5 pe assay kit

After transfection and incubation for 48 h, gastric cancer cells were stained with the Annexin V-PE assay kit (BD Biosciences, USA) based on the manufacturer’s instructions. All specimens were performed using the FACS Caliber II sorter and the Cell Quest FACS system (BD Biosciences, USA) for analysis.

The CCK-8 detection kit (Dojindo, Shanghai, China) was used to measure cell viability. Cells were seeded in a 96-well (100 μl per well) plates at a density of 5 × 10⁴/ml. CCK-8 solution (10 μl per well) was added and the plate was incubated at 37 °C for 2 h. The viable cells were counted by absorbance measurements with a monochromator microplate reader at a wavelength of 450 nm.
BrdU incorporation and cell proliferation analysis were detected using BD Pharmingen™ BrdU Flow Kit (BD, San Diego, CA, USA). Cells were plated into 6-well plates and transfected with siRNA for 24 h, added BrdU (1 mM/mL) for 12 h and then harvested. Cells were stained with anti-BrdU and 7-AAD according to the manufacturer’s instructions. Cell cycle distribution was determined by flow cytometry (FACSAriaTM II, Becton Dickinson, Mountain View, CA, USA).
The Annexin V-PE assay kit (BD) was utilized to analyze cell apoptosis in GBM cells. After transfected with siRNA for 24 h, cells were harvested and resuspended in 100 ul binding buffer at a density of 1 × 10⁶ cells/ml. PE-conjugated Annexin V and 7-AAD reagent staining were performed in concentrations and time recommended by the manufacturer. Stained cells were analyzed by flow cytometry (FACSAria II, BD).