Advanced fluorescence lifetime-enhanced multiplexed nanoscopy of cells.

In this review paper, we summarize the significant advancements in the field of fluorescence lifetime imaging microscopy (FLIM), particularly wide-field FLIM with single-molecule sensitivity, achieved using the time-correlated single-photon counting-based position-sensitive LINCam system. Fluorescence lifetime adds valuable information beyond conventional intensity-based imaging, enabling diverse applications across research fields. Here, we focus on three primary bioimaging applications: (I) single-molecule FLIM in the far-red spectral region, (II) fast and multiplexed super-resolution imaging of cells, and (III) three-dimensional super-resolution imaging with high axial localization precision. Recent advances in position-sensitive detector technologies offer exciting opportunities for high-throughput super-resolution imaging with enhanced localization precision.