Single pixel imaging frameworks facilitate the acquisition of high-dimensional optical data in biological applications with photon starved conditions. However, they are still limited to slow acquisition times and low pixel resolution. Herein, we propose a convolutional neural network for fluorescence lifetime imaging with compressed sensing at high compression (NetFLICS-CR), which enables in vivo applications at enhanced resolution, acquisition and processing speeds, without the need for experimental training datasets. NetFLICS-CR produces intensity and lifetime reconstructions at 128 × 128 pixel resolution over 16 spectral channels while using only up to 1% of the required measurements, therefore reducing acquisition times from ∼2.5 hours at 50% compression to ∼3 minutes at 99% compression. Its potential is demonstrated in silico, in vitro and for mice in vivo through the monitoring of receptor-ligand interactions in liver and bladder and further imaging of intracellular delivery of the clinical drug Trastuzumab to HER2-positive breast tumor xenografts. The data acquisition time and resolution improvement through NetFLICS-CR, facilitate the translation of single pixel macroscopic flurorescence lifetime imaging (SP-MFLI) for in vivo monitoring of lifetime properties and drug uptake.
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