Fig. 1. (a) Schematic of AO-TP-FLIM. HWP, half-wave plate; PBS, polarization beam splitter; M, mirror; L, lens; SLM, spatial light modulator; DM, dichroic mirror; OB, object lens; F, bandpass filter; PMT, photomultiplier tube; FPGA, field programmable gate array; TCSPC, time-correlated single-photon counting.

Fig. 2. Calibration of system resolution. (a) The pink box is the imaging of fluorescent beads before AO correction, and the blue box is the imaging of fluorescent beads after AO correction. (b) System resolution before and after correction (normalized); (c) calibration of fluorescence lifetime and time resolution of AO-TP-FLIM microscopic imaging system. The yellow box (e) is the fluorescence lifetime imaging of sodium fluorescein before AO correction, and the green box (f) is the fluorescence lifetime imaging of sodium fluorescein after AO correction. (d) Comparison of fluorescence lifetime histograms of sodium fluorescein before and after AO correction. The scale is 0.2 µm.

Fig. 3. Fluorescent bead imaging data analysis under 450 µm mice brain tissue. (a) Fluorescence lifetime imaging before AO correction; (b) fluorescence lifetime imaging after AO correction; (c) phase vector map before AO correction; (d) phase vector map after AO correction; (e) fluorescence intensity before and after AO correction (the inset is the phase diagram). (f) Fluorescence lifetime histogram before and after AO correction. The scale is 2 µm.

Fig. 4. Imaging of Aβ plaques in different areas of AD mice. (a) Fluorescence lifetime imaging and phase vector imaging before AO correction, with orange and blue box positions representing plaque areas outside and inside blood vessels, respectively; (b) fluorescence lifetime imaging and phase vector map after AO correction, with pink and green box positions representing the plaque areas outside and inside the blood vessels, respectively; (c) fluorescence intensity of extravascular Aβ plaques before and after AO correction (the inset is the phase diagram); (d) fluorescence intensity of intravascular Aβ plaques before and after AO correction; (e) fluorescence lifetime histogram of extravascular plaques before and after AO correction; (f) fluorescence lifetime histogram of intravascular plaques before and after AO correction. The scale is 10 µm.

Fig. 5. In vivo imaging of transgenic zebrafish larvae. (a) Fluorescence lifetime imaging before AO correction; (b) fluorescence lifetime imaging after AO correction; (c) phase vector map before AO correction; (d) phase vector map after AO correction; (e) fluorescence intensity before and after AO correction (the inset is the phase diagram); (f) fluorescence lifetime histogram before and after AO correction. The scale is 10 µm.