Title: Computational Dead Time Correction in Fluorescence Lifetime Imaging Microscopy (FLIM)

Abstract: Fluorescence Lifetime Imaging Microscopy (FLIM) is a crucial tool in biomedical research. It helps visualize and measure molecular interactions and cellular processes by analyzing the fluorescence decay of molecules in biological samples. Unlike traditional imaging methods based on fluorescence intensity, FLIM uses fluorescence lifetime - the time delay between excitation and emission in the fluorescence process - to provide detailed information about the biochemical environment and molecular activity.

In photon-counting systems, dead time is the short period after detecting a photon when the detection system cannot record new photons. The effect of dead time on the measured fluorescence intensity and lifetime is highly dependent on the photon rate (the number of photons detected per laser period). Dead time causes photon counts to be underestimated, especially at high photon rates, leading to errors in both fluorescence intensity and lifetime measurements.

Here, we developed a computational algorithm to correct for dead time effects in FLIM data. Using methods inspired by Light Detection and Ranging (LiDAR) systems, we improved the accuracy of FLIM results through statistical modeling, simulations, and evaluation with simulated FLIM data.