Comparison of the penetration depth in mouse brain in vivo through 3PF imaging using AIE nanoparticle labeling and THG imaging within the 1700 nm window.

3-Photon microscopy (3PM) excited at the 1700 nm window features a smaller tissue attenuation and hence a larger penetration depth in brain imaging compared with other excitation wavelengths in vivo . While the comparison of the penetration depth quantified by effective attenuation length l e with other excitation wavelengths have been extensively investigated, comparison within the 1700 nm window has never been demonstrated. This is mainly due to the lack of a proper excitation laser source and characterization of the in vivo emission properties of fluorescent labels within this window. Herein, we demonstrate detailed measurements and comparison of l e through the 3-photon imaging of the mouse brain in vivo , at different excitation wavelengths (1600 nm, 1700 nm, and 1800 nm). 3PF imaging and in vivo spectrum measurements were performed using AIE nanoparticle labeling. Our results show that l e derived from both 3PF imaging and THG imaging is the largest at 1700 nm, indicating that it enables the deepest penetration in brain imaging in vivo .