High Speed Spectral Fluorescence Lifetime Imaging for Lifescience Applications

摘要

We present an achromatic confocal laser scanning system capable of recording spectrally resolved fluorescence lifetimeimages (sFLIMs) at a rate of >8 frames per second (FPS) for a 128 x 128 image. This frame rate was achieved byoptimizing the processing of lifetime calculations from previous results which demonstrated >4 FPS sFLIM imaging.The imaging system is achromatic for a spectral range of 400 - 900 nm, achieved by using reflective optics instead of atransmissive lens system, except for the primary objectives. Two excitation sources have been integrated into the system,485 nm and 640 nm laser diodes with a pulse width of <70 ps and <90 ps respectively. Imaging is performed via agalvanometric mirror system which scans the laser beam over the sample with the ability to change the Field of View(FOV) on the fly. The collected fluorescence signal is focused into a multimode fiber via a second objective andrecollimated onto a transmissive grating for spectral dispersion onto a novel complementary metal–oxide–semiconductorsingle photon avalanche diode (CMOS SPAD) line array sensor. This sensor can perform lifetime histogram generationon-chip and process over 16.5 Giga events/s enabling fast lifetime data acquisition. High speed sFLIM is demonstratedthrough imaging of convallaria majalis sections.