Description
Key facts
Sample types:Suspension cells, Adherent cells,
Assay type:Cell-based,
Assay time:1h,
Assay Platform:Flow cytometer, Fluorescence microscope
Product details:
Apoptosis/ Necrosis Assay Kit (blue, green, red) ab176749 is designed to simultaneously stain apoptotic (green), necrotic (red) and healthy cells (blue). Analysis is by flow cytometer or fluorescence microscope.
How the assay works
Apoptotic cells are stained using a fluorescent sensor for phosphotidylserine (PS); Apopxin Green Indicator. The PS sensor used has green fluorescence (Ex/Em = 490/525 nm) upon binding to membrane PS.
Cells with loss of plasma membrane integrity, ie late stage apoptotic and necrotic cells, are stained with the membrane-impermeable 7-AAD (Ex/Em = 546/647 nm), which is used to label the nucleus.
Live cells are stained with the live cell cytoplasm labeling reagent, CytoCalcein Violet 450. This reagent is cell permeable, and after entering live cells is hydrolyzed into a fluorescent dye (Ex/Em = 405/450 nm).
Apoptosis/ Necrosis assay protocol summary
- Centrifuge cells and resuspend in assay buffer
- Add Apopxin Green, 7-AAD and CytoCalcein Violet 450
- Incubate at room temperature for 30 - 60 minutes
- Analyze cells with a flow cytometer or fluorescence microscope
Related and recommended products
Alternative kits to monitor apoptosis/ necrosis are available:
- Apoptosis/ Necrosis Assay Kit (green, orange, red)
ab270781
- Apoptosis/ Necrosis Assay Kit (blue, red, green)
ab176750
Other Notes
This product was previously called Apoptosis/ Necrosis Detection Kit.
Properties and Storage Information:
Shipped at conditions-Blue Ice, Appropriate short-term storage conditions--20°C, Appropriate long-term storage conditions--20°C, Storage information--20°C
Supplementary Information:
This supplementary information is collated from multiple sources and compiled automatically.
Apoptosis and necrosis are distinct forms of cell death with mechanical and biological differences. Apoptosis is a highly regulated process often called programmed cell death while necrosis is an uncontrolled cell death resulting from injury or infection. Apoptosis involves a series of signaling pathways including the activation of caspases a family of proteases that play essential roles in cell death. The apoptotic process takes place in various tissues and is important for maintaining cellular homeostasis. Necrosis on the other hand typically occurs due to acute damage and is characterized by cell swelling membrane rupture and inflammation. These processes can be investigated using tools like apoptosis detection kits and necrosis markers.
Biological function summary
Apoptosis serves to eliminate damaged or unnecessary cells without eliciting an inflammatory response. It is part of the intrinsic and extrinsic pathways that incorporate various proteins and signaling molecules. During apoptosis the formation of complexes such as the apoptosome triggers caspase activation. Key players in apoptosis also include Bcl-2 family proteins which regulate mitochondrial membrane permeability. Conversely necrosis results in a direct inflammatory response due to the release of intracellular contents. Both apoptosis and necrosis play roles in developmental processes and maintaining tissue integrity.
Pathways
The apoptotic process integrates into pathways like the mitochondrial (intrinsic) pathway and the death receptor (extrinsic) pathway. In the intrinsic pathway cytochrome c release from mitochondria forms the apoptosome complex with Apaf-1 and caspase-9 leading to downstream caspase activation. The extrinsic pathway involves death receptors such as Fas and TNF receptor interacting with adaptor proteins to activate caspase-8. Necrosis typically results from pathophysiological conditions but can share overlaps with necroptosis a regulated form of necrosis involving proteins such as RIPK1 and RIPK3.
Dysregulation of apoptosis and necrosis is linked to conditions like cancer and neurodegenerative diseases. Abnormal apoptosis is associated with cancer where reduced apoptosis allows uncontrolled cell proliferation. This often involves proteins such as p53 which can promote apoptosis under stress conditions but mutations can alter its function. In neurodegenerative diseases like Alzheimer's both apoptosis and necrosis contribute to neuronal loss. The accumulation of misfolded proteins like amyloid-beta may trigger apoptotic pathways while necrosis can result from oxidative stress and excitotoxicity.
