Fig. 1. Schematic illustration of 2PFLIM system (PMT as the detector) in the NIR-II region. 0) 1550 nm fs laser; 1) half-wave plate; 2) polarization beam splitter (PBS); 3) photodiode; 4,5) reflector; 6) scanning galvanometer; 7,8,11) lens; 9) dichroic mirror; 10) objective; 12) large beam collimator; 13) PMT; 14) TCSPC board; 15) computer. Dashed box is the scanning microscope.

Fig. 2. Morphological characterization and optical properties of PbS/CdS quantum dots (PbS QDs). (a) Transmission electron microscopy (TEM) image of PbS QDs. (b) Dynamic light scattering (DLS) profile of PbS QDs. (c) One-photon and multiphoton fluorescence spectra of PbS QDs. (d) Relationship of 1550 nm fs excitation power and fluorescence intensity from QDs.

Fig. 3. In vivo 2PFLIM images of QDs-stained mouse brain blood vessels with PMT. (a) 2PF intensity images of brain blood vessels at different vertical penetration depths: from 0 to 220 μm. Scale bar: 50 μm. (b) 2PF lifetime image at 60 μm. (c) Time-resolved decay curve at the arrow in (b). (d) 2PFLIM image at 220 μm. (e) Time-resolved decay curve at the arrow in (d). (f)–(i) Plots of each FWHM of blood vessels at depth of 100, 140, 180, and 220 μm, as indicated by the yellow lines in (a), respectively.

Fig. 4. (a) Schematic illustration of 2PFM system (InGaAs camera as the detector) in the NIR-II region. 0) 1550 nm fs laser; 1) half-wave plate; 2) polarization beam splitter (PBS); 3,4) reflector; 5) scanning galvanometer; 6,7,10) lens; 8) dichroic mirror; 9) objective; 11) InGaAs camera; 12) computer. Dashed box is the scanning microscope. (b) 3D reconstructed image of fluorescence spots when laser is focused at different depths. (c) Statistics of fluorescence area at various depths.

Fig. 5. Schematic diagram of image overlay: the blood vessel image in the blue box is superimposed by the other 15 frames. Scale bar: 50 μm.

Fig. 6. In vivo 2PFM images of QDs-stained mouse brain blood vessels with InGaAs camera. (a) 2PFM images of brain blood vessels at different vertical penetration depths: from 0 to 110 μm. Scale bar: 50 μm. (b)–(d) Plots of each FWHM of blood vessels at depths of 40, 60, and 80 μm as indicated by the yellow lines in (a), respectively.