Principle of Assay
Apoptosis is a form of programmed cell death to remove unwanted, damaged, or senescent cells from tissues. In normal cells, the negative phospholipids reside on the inner side of the cellular membrane while the outer surface of the membrane is occupied by uncharged phospholipids (PS). After a cell has entered apoptosis, the negatively charged PS are transported from the inner to the outer leaflet of the plasma membrane, thus exposing PS to the external cellular environment. The human anticoagulant, Annexin V, is a 35-36 kDa Ca2+ dependent phospholipid-binding protein that has a high affinity for PS. Annexin V labeled with a fluorophore or biotin can identify apoptotic cells by binding to PS exposed on the outer leaflet. Propidium iodide (PI) is a fluorescent nucleus dye, impermeant to live cells and apoptotic cells, but stains dead cells with red fluorescence, binding tightly to the nucleic acids in the cell. Annexin V-EGFP/PI Apoptosis Detection kit provides a rapid and convenient assay for apoptosis. After staining a cell population with Annexin V-EGFP/PI in the provided binding buffer, early apoptotic cells show green fluorescence of the cellular membrane, dead cells show red fluorescence of the nucleus and green fluorescence of the cellular membrane, and live cells show little or no fluorescence. In addition, to address the non-standardization of apoptosis detection and results, the kit also provides a patented positive reference substance for apoptosis. Detection can be analyzed by flow cytometry or by fluorescence microscopy.
