Annexin 5 kit

Test of cell apoptosis was via exerting Annexin V Kit (Beyotime, Beijing, China). Detachment of the cells was with 0.25% trypsin, removal of digestive juice was conducted, and placing of the cells was in the previously collected medium. Centrifugation at 12,000 RPM and abandonment of the supernatant were performed. Collection of the cell precipitates and production of the cell suspension with phosphate buffer saline were conducted. Centrifugation again and removal of the supernatant were conducted; Addition of Annexin V fluorescein-isothiocyanate and 300 mL propidium iodide labeling solution and incubation were performed [7 (link)].

The Annexin V kit (Beyotime Institute of Biotechnology, Beijing, China) was used to detect cell apoptosis. Twelve h after HeLa, SiHa, and C33A cells received different treatments as described above, they were exposed to 0, 2, 4, 6, or 8 Gy doses of X-rays. The cells were then cultured for another 48 h, transferred to centrifuge tubes, washed with PBS, and digested with 0.25% trypsin. The digestion solution was then removed and replaced with the original culture solution. The cells were centrifuged at 12000 rpm for 5 min at 4°C, the supernatant was discarded, cell pellets were resuspended (5-10×10⁴U) in PBS solution, centrifuged again for 5 minutes, and the supernatant was discarded. The cells were then resuspended with 200 μL of Annexin V-FITC and incubated in the dark at room temperature for 10 min. The cells were then centrifuged again for 5 min, the supernatant was discarded, and 190 μL of Annexin V-FITC and 10 μL of propidium iodide (PI) was added. The cells were then mixed well and placed in an ice bath. Cell apoptosis was analyzed using a FACSCanto II (Becton-Dickinson, Franklin Lakes, NJ, USA). Apoptosis rates were calculated as early apoptosis rate + late apoptosis rate.

Cell apoptosis was detected by the Annexin V Kit (Beyotime, Beijing, China). Cells were digested with 0.25% trypsin, and the digestion fluid was removed and cells were placed into the previously collected culture medium. After centrifuging at 12,000 RPM for 5 min at 4°C, the supernatant was discarded. Cell precipitation was collected to make cell suspensions using PBS. After another centrifugation and supernatant removal, 300 ml of Annexin VFITC and propidium iodide-labeled liquid were added to culture the cells at 4°C in the dark for 30 min. The flow cytometry (BD Pharmingen, San Diego, CA, USA) was used to detect cell apoptosis.