Flow cytometry (BD FACSAria; BD Biosciences, Franklin Lakes, NJ, USA) was performed to detect cell apoptosis. The following 2 groups were under investigation: i) the control group and ii) the 1% oxygen group. We observed the apoptotic rates at 48 h post-treatment. In accordance with the Annexin V/propidium iodide (PI) apoptosis kit (BioVision, San Francisco, CA, USA), 5×105 cells were collected in each tube and 1 ml Annexin V binding buffer was added followed by thorough mixing. Subsequently, 5 µl Annexin V-fluorescein isothiocyanate and 10 µl PI were added. After mixing, the tube was incubated in the dark at 37°C for 15 min. For the early apoptotic cells, membrane phosphatidylserine was exposed and combined with Annexin V but no PI. For the late apoptotic cells, the membranes were permeable to PI and the cells were stained with Annexin V and PI. The dead cells were stained only with PI.
Annexin 5 propidium iodide apoptosis kit
Manufactured by Abcam
Sourced in United States
Annexin V/propidium iodide (PI) apoptosis kit is a laboratory tool used to detect and quantify apoptosis, a form of programmed cell death. The kit contains Annexin V, a protein that binds to phosphatidylserine, and propidium iodide, a DNA-binding dye. These components are used in flow cytometry or fluorescence microscopy to distinguish between viable, early apoptotic, and late apoptotic/necrotic cells.
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4 protocols using annexin 5 propidium iodide apoptosis kit
1
Apoptosis Analysis by Flow Cytometry
2
Annexin V-FITC/PI Apoptosis Assay
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Five groups as mentioned earlier (see “Chemicals and Treatments” section) were investigated. Apoptotic rates were examined 48 h after the treatment. In accordance with the Annexin V/Propidium Iodide (PI) Apoptosis Kit (BioVision, CA, USA), 5 × 105 cells were collected in each tube and 1 ml of annexin V binding buffer was added, followed by thorough mixing. Subsequently, 5 ml of annexin V–fluorescein isothiocyanate and 10 ml of PI were added. After mixing, the tube was incubated in the dark at 37°C for 15 min. For the early apoptotic cells, the membrane phosphatidylserine was exposed and combined with annexin V, without PI. For the late apoptotic cells, the membranes were permeable to PI and the cells were stained with annexin V and PI. The dead cells were stained only with PI. The samples were analyzed using a FACScan flow cytometer (BD Biosciences, NJ, USA) within 1 h. Flow cytometry (BD FACSAria; BD Biosciences) was performed to detect cell apoptosis.
3
Apoptosis Detection by Annexin V/PI
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Cell apoptosis was detected in accordance with the Annexin V/propidium iodide (PI) apoptosis Kit (BioVision, U.S.A.). In brief, 4 × 105 cells were added in each tube. Subsequently, 5 µl Annexin V-fluorescein isothiocyanate and 10 µl PI were added. After mixing, the tube was incubated at 37°C for 15 min in the dark. Analysis was performed using a FACSCalibur flow cytometer.
4
Apoptosis Assessment via Flow Cytometry
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The following 5 groups mentioned above (see "Experimental design and ischemic injury cell Model" section) were under investigation. Apoptotic rates were examined 48 h post-treatment. In accordance with the Annexin V/propidium iodide (PI) apoptosis kit (BioVision, San Francisco, CA, USA), 5 × 10 5 cells were collected in each tube and 1 mL Annexin V binding buffer was added followed by thorough mixing. Subsequently, 5 μL Annexin V-fluorescein isothiocyanate and 10 μL PI were added. After mixing, the tube was incubated in the dark at 37 °C for 15 min. For the early apoptotic cells, membrane phosphatidylserine was exposed and combined with Annexin V, without PI. For the late apoptotic cells, the membranes were permeable to PI and the cells were stained with Annexin V and PI. The dead cells were stained only with PI. The samples were analyzed using a FACScan flow cytometer (BD Biosciences, USA) within 1 h. Flow cytometry (BD FACSAria; BD Biosciences, Franklin Lakes, NJ, USA) was performed to detect cell apoptosis.
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