Key features and details
- Assay type: Quantitative
- Detection method: Fluorescent
- Platform: Flow cytometer
- Sample type: Adherent cells, Suspension cells
Product nameAnnexin V-iFluor 350 Apoptosis Staining / Detection Reagent
Sample typeAdherent cells, Suspension cells
Species reactivityReacts with: Mammals, Other species
Annexin V-iFluor 350 Apoptosis Staining / Detection Reagent (ab219903) is a cell-impermeable reagent designed to bind to phosphatidylserine (PS) residues exposed on the outer cell surface of cells with a flow cytometer or fluorescence microscopy at Ex/Em = 347/443 nm.
We recommend using an impermeable nuclear stain such as Propidium Iodide (ab14083) or DRAQ7™ (ab109202) together with Annexin V-iFluor 350 Detection Reagent to discriminate necrotic and dead cells: plasma membrane is disrupted in these cells and therefore the Annexin V reagent will bind to PS found in the interior of cells.
Apoptosis is a regulated process of cell death that occurs during embryonic development as well as maintenance of tissue homeostasis. Inappropriately regulated apoptosis is implicated in different disease states, such as neurodegeneration disease and cancer. The apoptosis program is characterized by morphologic features, including loss of plasma membrane asymmetry and attachment, condensation off the cytoplasm and nucleus, and compaction and fragmentation of the nuclear chromatin. Exposure of phosphatidylserine (PS) on the external surface of the cell membrane has been reported to occur in the early phases of apoptotic cell death, during which the cell membrane remains intact. In leukocyte apoptosis, PS on the outer surface of the cell marks the cell for recognition and phagocytosis by macrophages. The human vascular anticoagulant, annexin V, is a 35-36 kDa Ca2+ dependent phospholipid binding protein that has a high affinity for PS, and shows minimal binding to phosphatidylcholine and sphingomyelin. Changes in PS asymmetry, which can be analyzed by measuring annexin V binding to the cell membrane, are generally observed before morphological changes associated with apoptosis occurred and before membrane integrity is lost.
Storage instructionsStore at -20°C. Please refer to protocols.
Components 100 tests Annexin V-iFluor 350 conjugate 1 x 200µl
Annexin V-mFluor Violet450 Detection Reagent (ab219911) which has similar PS binding properties to Annexin V-iFluor 350 conjugate, Detection of phosphatidylserine (PS) exposure in Jurkat cells. Jurkat cells were left untreated (blue) or treated with 1 μM staurosporine (red) in a 37°C, 5% CO2 incubator for 5 hours. Cells were then incubated with Annexin V Detection Reagent for 30 minutes. The fluorescence intensity of Annexin V-mFluor 450 Detection Reagent was measured with a FACSCalibur (BD systems)flow cytometer using violet laser at Ex/Em = 405/450 nm.
In live non-apoptotic cells, Annexin V-iFluor 350 conjugate detects innate apoptosis in non-induced cells, which is typically 2-6% of all cells. In apoptotic cells Annexin V-iFluor 350 conjugate binds to phosphatidylserine, which is located on the outer leaflet of the cell membrane, resulted in increased staining intensity.