Scattering Correction through Fourier-Domain Intensity Coupling in Two-Photon Microscopy (2P-FOCUS)

Area of Science:

• Biomedical Optics
• Microscopy
• Biophysics

Background:

• Light penetration in biological tissues is limited by scattering.
• Correcting scattering is difficult with limited photons or without transmission access.

Purpose of the Study:

• Introduce a novel two-photon microscopy system with Fourier-domain intensity coupling for scattering correction (2P-FOCUS).
• Address challenges in deep tissue imaging caused by light scattering.

Main Methods:

• Utilize Fourier-domain intensity modulation and nonlinear interference for scattering correction.
• Employ random patterns and a single-shot algorithm for rapid mask generation.
• Overcome memory effect limitations by customizing correction masks for subregions.

Main Results:

• Demonstrated focusing and imaging through bone and mouse brain samples.
• Significantly enhanced two-photon fluorescence signals (tens of folds) compared to uncorrected imaging.
• Achieved scattering correction over a large volume (230×230×510 μm³), exceeding the memory effect range.
• Completed scattering measurement, calculation, and correction within seconds.

Conclusions:

• 2P-FOCUS effectively corrects scattering, enabling deeper light penetration into tissues.
• The system offers speed, effectiveness, and cost-efficiency for broad adoption in deep tissue imaging.
• 2P-FOCUS eliminates the need for guide stars, iterative optimization, or transmission/reflection measurements.