The RealTime-Glo™ Annexin V Apoptosis and Necrosis Assay measures the real-time exposure of phosphatidylserine (PS) on the outer leaflet of cell membranes during the apoptotic process. Annexin V luciferase fusion proteins supplied in the assay reagent bind to PS during early apoptosis and are detected with a simple luminescence signal. The reagent also includes a DNA-binding dye, which enters the cell and generates a fluorescent signal upon loss of membrane integrity.

The combination and timing of luminescent and fluorescent signals is used to differentiate secondary necrosis occurring during late apoptosis from necrosis caused by other cytotoxic events.

The assay is non-lytic and the simple “add-and-read” method allows multiple readings from a single assay well. Apoptosis can be monitored in real time, without the need for multiple plates, complicated processing, or specialized detection equipment. A multimode reader capable of detecting luminescence and fluorescence is the only instrument required.

Based on NanoLuc® Binary Technology (NanoBiT), the Realtime-Glo™ Annexin V Apoptosis Assay uses Annexin V fusion proteins containing complementary subunits of NanoBiT® luciferase (Annexin V-LgBiT and Annexin V-SmBiT). When in close proximity, the LgBiT and SmBiT subunits come together to form a functional luciferase.

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