Adaptive optical third-harmonic generation microscopy for in vivo imaging of tissues

Cristina Rodríguez; Daisong Pan; Ryan G Natan; Manuel A Mohr; Max Miao; Xiaoke Chen; Trent R Northen; John P Vogel; Na Ji

doi:10.1101/2024.05.02.592275

Adaptive optical third-harmonic generation microscopy for in vivo imaging of tissues

bioRxiv [Preprint]. 2024 May 5:2024.05.02.592275. doi: 10.1101/2024.05.02.592275.

Authors

Cristina Rodríguez^{1

2

3}, Daisong Pan¹, Ryan G Natan^{1

2}, Manuel A Mohr^{4

5}, Max Miao^{6

7}, Xiaoke Chen⁴, Trent R Northen^{6

7}, John P Vogel^{6

7}, Na Ji^{1

2

8

9

10}

Affiliations

¹ Department of Physics, University of California, Berkeley, CA, USA.
² Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, VA, USA.
³ Present address: Department of Biomedical Engineering, Yale University, New Haven, CT, USA.
⁴ Department of Biology, Stanford University, Stanford, CA, USA.
⁵ Present address: Yale Ventures, Yale University, New Haven, CT, USA.
⁶ Environmental Genomics and Systems Biology, Lawrence Berkeley National Laboratory, Berkeley, CA, USA.
⁷ Department of Energy Joint Genome Institute, Lawrence Berkeley National Laboratory, Berkeley, CA, USA.
⁸ Department of Molecular and Cell Biology, University of California, Berkeley, CA, USA.
⁹ Helen Wills Neuroscience Institute, University of California, Berkeley, CA, USA.
¹⁰ Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, CA, USA.

Abstract

Third-harmonic generation microscopy is a powerful label-free nonlinear imaging technique, providing essential information about structural characteristics of cells and tissues without requiring external labelling agents. In this work, we integrated a recently developed compact adaptive optics module into a third-harmonic generation microscope, to measure and correct for optical aberrations in complex tissues. Taking advantage of the high sensitivity of the third-harmonic generation process to material interfaces and thin membranes, along with the 1,300-nm excitation wavelength used here, our adaptive optical third-harmonic generation microscope enabled high-resolution in vivo imaging within highly scattering biological model systems. Examples include imaging of myelinated axons and vascular structures within the mouse spinal cord and deep cortical layers of the mouse brain, along with imaging of key anatomical features in the roots of the model plant Brachypodium distachyon. In all instances, aberration correction led to significant enhancements in image quality.

Publication types

Preprint

Grants and funding

U01 NS118300/NS/NINDS NIH HHS/United States