# Computational two-photon microscopy for deep tissue imaging

> **NIH NIH R35** · UNIVERSITY OF CALIFORNIA AT DAVIS · 2024 · $374,399

## Abstract

ABSTRACT
Intravital microscopy has been widely employed to visualize the structures and functions of cells in their natural
environment. However, the imaging depth of intravital imaging is constrained by tissue scattering. State-of-the-
art multiphoton microscopy can only produce sharp images up to about 1 mm deep in biological tissues at cellular
resolution, which restricts numerous biological applications requiring access deep inside tissues. Deep tissue
imaging faces two major challenges. First, the 3D refractive index of tissues, often referred to as the 'scattering
potential,' undergoes dynamic changes in living organisms. Second, the total light power for illuminating
biological tissue is limited to prevent thermal damage. Therefore, optical imaging techniques that are capable of
high-speed scattering correction and maximizing the light power at the focal plane are needed.
In this proposal, our group aims to develop innovative two-photon microscopy systems with scattering correction
for deep tissue imaging. The fundamental concept behind our techniques is coupling the illumination light into
high-transmission channels (referred to as 'open channels') within the scattering tissues while mitigating light in
the low-transmission channels. To achieve this goal, we will employ various techniques, encompassing
advancements in optical hardware and computational software. The hardware innovation is to develop high-
speed, high-efficiency light modulation, while the software innovation is to develop new deep-learning-based
algorithms for optimizing correction patterns and reducing computational complexity. Our proof-of-concept
experiments demonstrated a remarkable 27-fold increase in fluorescent intensity for imaging through a bone
sample after scattering correction. Our goal is to complete the entire correction process within 10 ms and achieve
a 100-fold increase in fluorescent signal after correction. We will validate our techniques by imaging intact mouse
organs. We will assess our techniques by comparing the image signal-to-noise ratio and the resolution with the
ground truth images obtained after tissue sectioning.
The novel two-photon microscopy techniques with scattering correction will enable deep tissue imaging at
previously unattainable depths while achieving cellular resolution. In the long term, we will collaborate with my
colleagues in the Alzheimer’s Disease Research Center and the Comprehensive Cancer Center at UC Davis to
apply the microscopy to broad biological research. This technique will facilitate the study of cellular structures
and activity within living organisms, such as neuronal activity deep in the brain, immune responses in the lymph
nodes, cancer metastasis, and hematopoietic stem cells in the bone marrow.

## Key facts

- **NIH application ID:** 10941812
- **Project number:** 1R35GM155193-01
- **Recipient organization:** UNIVERSITY OF CALIFORNIA AT DAVIS
- **Principal Investigator:** Yi Xue
- **Activity code:** R35 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $374,399
- **Award type:** 1
- **Project period:** 2024-08-01 → 2029-07-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10941812

## Citation

> US National Institutes of Health, RePORTER application 10941812, Computational two-photon microscopy for deep tissue imaging (1R35GM155193-01). Retrieved via AI Analytics 2026-05-25 from https://api.ai-analytics.org/grant/nih/10941812. Licensed CC0.

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