Product Introduction:

Early apoptotic changes occur on the surface of the cell membrane, and one of these changes on the surface of the cell membrane is the transfer of phosphatidylserine (PS) from inside the cell membrane to outside the cell membrane, exposing PS to the outer surface of the cell membrane. PS is a negatively charged phospholipid, which normally exists in the inner side of the cell membrane. During cell apoptosis, the asymmetry of this phospholipid distribution on the cell membrane is destroyed, and PS is exposed to the outside of the cell membrane. Annexin V has easy binding to phospholipids such as PS and has a high affinity for PS. Thus, the protein can act as a sensitive probe to detect PS exposed on the surface of the cell membrane. The transfer of PS to the outside of the cell membrane is not unique to apoptosis and can also occur in cell necrosis. The difference between the two modes of cell death is that the membrane is intact in the initial stage of apoptosis, while the integrity of the membrane is destroyed in the early stages of cell necrosis. Therefore, the AnnexinV and PI double staining method can be used to detect the early apoptosis of cells by flow cytometry.

Operation steps :

labeled Annexin V can be incorporated into flow cytometry for the detection of phosphoacylserine on the outer membrane

1. Dilute the binding buffer with deionized water at 1:4 (4ml binding buffer +12ml deionized water);

2. The cells were washed twice with PBS pre-cooled at 4℃, and the cells were re-suspended with 250 UL combined buffer, and the concentration was adjusted to 1× 106/ml;

3. 100μl cell suspension was taken into a 5ml flow tube and 5μl Annexin V/Alexa Fluor 647 and 10μl 20ug/ml propyl iodide solution were added.

4. Mix well and incubate at room temperature for 15 minutes away from light;

5. 400μl PBS was added to the reaction tube for FACS analysis.

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