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Scattering correction through Fourier-domain intensity coupling in two-photon microscopy (2P-FOCUS)
Daniel Zepeda1, Yucheng Li1, Yi Xue1
1Department of Biomedical Engineering, University of California, Davis, 451 Health Sciences Dr., Davis, California 95616, USA.
Photonics Research
|July 4, 2025
Summary
We developed a new two-photon microscopy system (2P-FOCUS) that corrects light scattering in biological tissues. This system enhances deep tissue imaging by modulating light intensity in the Fourier domain, improving signal by tens of folds.
Area of Science:
- Biomedical optics
- Microscopy
- Photonics
Background:
- Light scattering in biological tissues limits imaging depth.
- Current scattering correction methods require transmission access or iterative optimization.
- Limited photon availability exacerbates scattering challenges.
Purpose of the Study:
- Introduce a novel two-photon microscopy system for scattering correction.
- Overcome limitations of existing scattering correction techniques.
- Enable deep tissue imaging in challenging conditions.
Main Methods:
- Developed a two-photon microscopy system with Fourier-domain intensity coupling for scattering correction (2P-FOCUS).
- Utilized intensity modulation in the Fourier domain, leveraging nonlinear interference and two-photon excitation.
- Employed random patterns for scattering probing and a single-shot algorithm for rapid correction mask generation.
- Enabled subregion-specific mask customization to overcome memory effect limitations.
Main Results:
- Demonstrated focusing and imaging through bone and ex vivo mouse brain samples.
- Significantly enhanced two-photon fluorescence signals (tens of folds) compared to uncorrected imaging.
- Achieved scattering correction in a volume of 230 μm × 230 μm × 510 μm, exceeding the memory effect range.
- Completed scattering measurement, calculation, and correction within seconds.
Conclusions:
- 2P-FOCUS effectively corrects light scattering in biological tissues.
- The system enables rapid, cost-effective deep tissue imaging.
- 2P-FOCUS offers broad adoption potential for advanced biomedical imaging applications.
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