Labeled annexin-V, with its high affinity for phosphatidylserine (PS), is a sensitive probe for PS exposed on the outer layer of apoptotic cells. Since necrotic cells also expose PS as a result of lost membrane integrity, propidium iodide is utilized as a DNA stain to distinguish necrotic cells from annexin-V-labeled cell clusters. Other secondary labeling is possible, such as membrane-surface staining with phycoerythrin- or TRITC-labeled monoclonal antibodies for further cellular characterization.
In the early stages of apoptosis, changes occur at the cell surface. One of these plasma membrane alterations is the translocation of phosphatidylserine (PS) from the inner side of the plasma membrane to the outer layer, exposing PS at the surface of the cell. Macrophages specifically recognize PS exposed on the cell surface of lymphocytes during the development of apoptosis. The recognition and phagocytosis of apoptotic cells and bodies protects organisms from exposure to cellular compounds that lead to inflammation, which usually accompanies necrosis. Annexin V is a 35.8kDa, Ca2+ -dependent phospholipid-binding protein with a high affinity for PS, making labeled annexin an excellent detection agent.
Detection: Annexin-V-FLUOS can be directly detected in FACS analysis and immunochemistry without a secondary detection system.
Sample material: Cell lines and freshly isolated cells.
Kit with FLUOS-conjugated annexin-V and propidium iodide for the detection and quantification of apoptosis and differentiation from necrosis at the single-cell level.
