Phasor EO-FLIM: Lifetime imaging with picosecond noise and 500 Hz frame rate

Fluorescence lifetime microscopy is limited by photon throughput, constraining the speed and dynamic range of biological measurements. We demonstrate a compact optical module and fast phasor acquisition methods to image multi-exponential lifetimes at 500 Hz in vivo and with over 1010 photons per second in static tissues. Action potentials are captured in phasor plots, and lifetime contrast is revealed from autofluorescence and wheat germ agglutinin stain with 20 picosecond pixel noise.