Two-photon light-sheet nanoscopy by fluorescence fluctuation correlation analysis.
Advances in light-sheet microscopy have enabled the fast three-dimensional (3D) imaging of live cells and bulk specimens with low photodamage and phototoxicity. Combining light-sheet illumination with super-resolution imaging is expected to resolve subcellular structures. Actually, such kind of super-resolution light-sheet microscopy was recently demonstrated using a single-molecule localization algorithm. However, the imaging depth and temporal resolution of this method are limited owing to the requirements of precise single molecule localization and reconstruction. In this work, we present two-photon super-resolution light-sheet imaging via stochastic optical fluctuation imaging (2PLS-SOFI), which acquires high spatiotemporal resolution and excellent optical sectioning ability. 2PLS-SOFI is based on non-linear excitation of fluctuation/blinking probes using our recently developed fast two-photon three-axis digital scanned light-sheet microscope (2P3A-DSLM), which enables both deep penetration and thin sheet of light. Overall, 2PLS-SOFI demonstrates up to 3-fold spatial resolution enhancement compared with conventional two-photon light-sheet (2PLS) microscopy and about 40-fold temporal resolution enhancement compared with individual molecule localization-selective plane illumination microscopy (IML-SPIM). Therefore, 2PLS-SOFI is promising for 3D long-term, deep-tissue imaging with high spatiotemporal resolution.
State Key Laboratory of Membrane Biology, Biodynamic Optical Imaging Center (BIOPIC), School of Life Sciences, Peking University, Beijing 100871, P.R. China. sun_yujie@pku.edu.cn.
Xuanze
X
Chen
Weijian
W
Zong
Rongqin
R
Li
Zhiping
Z
Zeng
Jia
J
Zhao
Peng
P
Xi
Liangyi
L
Chen
Yujie
Y
Sun