In-depth polarisation resolved SHG microscopy in biological tissues using iterative wavefront optimisation.

Area of Science:

• Biomedical Optics
• Molecular Imaging
• Biophysics

Background:

• Polarized nonlinear microscopy, particularly Polarization Second Harmonic Generation (PSHG), is vital for studying collagen organization in tissues.
• SHG imaging depth is limited by scattering and wavefront aberrations, typically to ~100 micrometers.
• Maintaining polarization control is critical for quantitative PSHG measurements.

Purpose of the Study:

• To investigate the impact of optical depth penetration on polarization control quality in PSHG imaging.
• To develop and evaluate wavefront shaping techniques for enhancing PSHG signal quality at depth.
• To enable in-depth PSHG imaging in collagen-rich biological tissues.

Main Methods:

• Applied adaptive optics for shallow-depth aberration correction.
• Utilized wavefront shaping of higher spatial frequencies for deeper focus correction.
• Employed nonlinear SHG active nanocrystals as guide stars for optimization.
• Tested methods on tendon and mouse cranial bone tissues.

Main Results:

• Demonstrated successful correction of wavefront aberrations and scattering effects.
• Quantified the recovery of high-quality linear polarization at increased depths.
• Showcased the potential for significantly improved in-depth PSHG imaging.

Conclusions:

• Wavefront shaping effectively restores polarization control for deep-tissue PSHG imaging.
• This approach overcomes current depth limitations in collagen organization studies.
• Enables advanced quantitative molecular analysis in intact biological tissues.