Apoptosis and necrosis are two major processes by which cells die. Apoptosis is an active, genetically regulated disassembly of the cell from within. During apoptosis, phosphatidylserine (PS) is translocated from the inner to the outer surface of the cell for phagocytic cell recognition. The human anticoagulant Annexin V is a 35 kDa, Ca2 -dependent phospholipid-binding protein with a high affinity for PS. Annexin V labeled with fluorescein (FITC) (λEx/λEm = 492/514 nm) can identify apoptotic cells in green by binding to PS exposed on the outer leaflet. Necrosis normally results from a severe cellular insult, resulting in loss of plasma membrane integrity. Ethidium homodimer III (EthD-III) is a highly positively charged nucleic acid probe, which is impermeant to live cells or apoptotic cells, but stains necrotic cells with red fluorescence (λEx/λEm (with DNA) = 528/617 nm). EthD-III is a superior alternative to propidium iodide (PI) or ethidium homodimer I because of its significantly higher affinity for DNA and higher fluorescence quantum yield. The kit also includes the membrane-permeant blue fluorescent DNA dye Hoechst 33342 (λEx/λEm (with DNA) = 350/461nm) for visualizing all cells. The kit allows simultaneous quantification of apoptotic (green), necrotic (red) and all cells (blue) within the same cell population by flow cytometry or fluorescence microscopy.
Kit Components:
- FITC-Annexin V
- Ethidium Homodimer III
- Hoechst 33342
- 5X binding Buffer
Features:
- Simple: Simultaneous detection of apoptotic, necrotic and healthy cells in a single assay
- Fast: A single 15-30 min staining step
- Sensitive: EthD-III is a superior alternative to both PI and EthD-I for staining necrotic cells.
Also see the Apoptosis and Necrosis Quantitation Kit Plus, and Apoptotic, Necrotic, and Healthy Cells Quantitation Kit Plus, which feature Annexin V conjugated to Biotium's superior green fluorescent dye CF®488A, which is brighter and more photostable than FITC.