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The detection principle of the Annexin V-Alexa Fluor488/PI apoptosis detection kit is as follows: in normal living cells, phosphotidylserine (PS) is located in the inner side of the cell membrane, but in early apoptotic cells, PS flips from the inner side of the cell membrane to the surface of the cell membrane and is exposed to the extracellular environment. Annexin-V is a Ca2+ -dependent phospholipid-binding protein with a molecular weight of 35 to 36KD, which binds with high affinity to PS and binds to the cell membrane of early apoptotic cells through phosphatidylserine exposed on the outside of the cell.
In addition, Propidium Iodide (PI) is provided to distinguish surviving early cells from necrotic or late apoptotic cells. PI is a nucleic acid dye, which can not pass through the intact cell membrane of normal cells or early apoptotic cells but can pass through the cell membrane of late apoptotic and necrotic cells and make the nucleus red. Thus, when Annexin V was combined with PI, PI was excluded from living cells (Annexin V-/PI-) and early apoptotic cells (Annexin V+/PI-). In contrast, late apoptotic and necrotic cells were double positive for Alexa Fluor488 and PI (Annexin V+/PI+)
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