Stroboscopic fluorescence lifetime imaging.

We report a fluorescence lifetime imaging technique that uses the time integrated response to a periodic optical excitation, eliminating the need for time resolution in detection. A Dirac pulse train of variable period is used to probe the frequency response of the total fluorescence per pulse leading to a frequency roll-off that is dependent on the relaxation rate of the fluorophores. The technique is validated by demonstrating wide-field, realtime, lifetime imaging of the endocytosis of inorganic quantum dots by a cancer cell line. Surface charging of the dots in the intra-cellular environment produces a switch in the fluorescence lifetime from approximately 40 ns to < 10 ns. A temporal resolution of half the excitation period is possible which in this instance is 15 ns. This stroboscopic technique offers lifetime based imaging at video rates with standard CCD cameras and has application in probing millisecond cell dynamics and in high throughput imaging assays.