Annexin 5 fluorescein isothiocyanate fitc cell apoptosis detection kit

After 48 h of transfection, the cells were detached with 0.25% trypsin without ethylene diamine tetraacetic acid (EDTA) (PYG0107, Boster, Wuhan, Hubei, China) and collected in a flow tube. The supernatant was discarded by centrifugation. According to the instructions of Annexin-V-fluorescein isothiocyanate (FITC) cell apoptosis detection kit (K201–100, BioVision, Palo Alto, USA), the Annexin-V-FITC, propidium iodide (PI), hydroxyethyl piperazine ethanesulfonic acid (HEPES) buffer solution was prepared into Annexin-V-FITC/PI staining solution at 1: 2: 50. Cells (1 × 10⁶) were resuspended in every 100 μL staining solution, then incubated for 15 min, and 1 mL HEPES buffer was added for mixing. The fluorescence of FITC and PI was detected by 515 nm and 620 nm band pass filters at 488 nm wavelength, and the apoptosis was detected.

At 48 h post‐transfection, the cells were trypsinized and dispersed into the cell suspension at 1 × 10⁶ cell/mL. A total of 1 mL suspension was centrifuged at 1500 r/min for 10 min and centrifuged again with 2 mL PBS. Subsequently, pre‐chilled 70% ethanol solution was added for overnight fixation of cells at 4℃. A total of 100 μL of the cell suspension (1 × 10⁶ cell/mL) reacted with 1 mL 50 mg/L propidium iodide (PI) containing RNAase for 30 min under conditions devoid of light and filtered. A flow cytometer was used to document the cell cycle detection under red fluorescence at the excitation wavelength of 488 nm.
The cells were trypsinized and centrifuged at 800 r/min for 5 min. According to the provided protocol of the Annexin V‐fluorescein isothiocyanate (FITC) cell apoptosis detection kit (K201‐100, BioVision, Inc., Exton, PA, USA), the cells were resuspended using 200 µL binding buffer and incubated with 10 µL PI staining solution and 5 µL Annexin V‐FITC for 15 min at room temperature under conditions devoid of light. Finally, a flow cytometer (Becton, Dickinson and Company, Franklin, NJ, USA) was used to detect the DNA content of cells at a wavelength of 488 nm.