Yong

Now you see me: A fluorogenic probe for cell surface phosphatidylserine enables time‐lapse imaging of cell death. The probe circumvents a number of drawbacks associated with commonly used annexin V cell death markers, including Ca²⁺ dependence, temperature sensitivity, and binding kinetics.

Abstract

The detection of externalized phosphatidylserine (PS) on the cell surface is commonly used to distinguish between living, apoptotic, and necrotic cells. The tools of choice for many researchers to study apoptosis are annexin V‐fluorophore conjugates. However, the use of this 35 kDa protein is associated with several drawbacks, including temperature sensitivity, Ca²⁺ dependence, and slow binding kinetics. Herein, a fluorogenic probe for cell surface PS, P‐IID, is described, which operates by an intramolecular indicator displacement (IID) mechanism. An intramolecularly bound coumarin indicator is released in the presence of cell surface PS, leading to a fluorescence “turn‐on” response. P‐IID demonstrates superior performance when compared to annexin V, for both fluorescence imaging and flow cytometry. This allows P‐IID to be used in time‐lapse imaging of apoptosis using confocal laser scanning microscopy and demonstrates the utility of the IID mechanism in live cells.