Annexin 5 fitc propidium iodide apoptosis detection kit

The tissue and cell suspension were transferred to a 15 mL centrifuge tube. After 5 minutes of centrifugation, PBS buffer was added to resuspend the tissue and cells. The apoptosis rate was detected using an Annexin V-FITC/propidium iodide (PI) apoptosis detection kit (Multisciences, Shanghai, China). After washing with ice-cold PBS for three times, cells were resuspended and incubated with 5 µL of Annexin V-FIFC and 10 µL of PI. A flow cytometer (BD Biosciences, NY, USA) was used to analyze cell apoptosis. The data were analyzed with FlowJo (Tree Star, OR, USA).

CasKi cells with 90% confluency were twice subjected to rinsing with phosphate-buffered saline (PBS) buffer with trypsin added to the culture, and then placed in a 37°C incubator for 3 min. After the cell synapses had disappeared, endothelial cell growth medium (EGM)-2 (Beijing Biterbo Biotechnology Co., Ltd., Beijing, China) containing 10% FBS was added to terminate the digestion. The medium was aspirated with a pipette, and the bottom of the culture dish was repeatedly blown on to promote cell shedding. Transfer of the cell suspension to a 15-mL centrifuge tube was followed by centrifugation for 5 min. Then, the tissue and cells were resuspended by adding PBS buffer. An annexin V-FITC/propidium iodide (PI) apoptosis detection kit (Multisciences, Shanghai, China) was selected for detection of the apoptotic rate. Cells were washed three times with ice-cold PBS before resuspension and incubation with 5 µL of Annexin V-FIFC and 10 µL of PI. A FACSAria Cell Sorter (BD Biosciences, Franklin Lakes, NJ, USA) was used for sample analysis. Data analyses were performed in Flow Jo (Tree Star, Ashland, OR, USA).