Multiline Orthogonal Scanning Temporal Focusing (mosTF) Microscopy for Scattering Reduction in High-speed in vivo Brain Imaging

TL;DR

This paper introduces mosTF microscopy, a novel imaging technique that significantly reduces scattering and improves image quality in high-speed in vivo brain imaging using temporal focusing two-photon microscopy.

Contribution

The paper presents mosTF, a new method that reconstructs scattered photons to enhance depth imaging in temporal focusing microscopy, overcoming previous limitations.

Findings

Improved signal-to-background ratio at greater depths

Enhanced resolution and image clarity in vivo

Maintained high-speed imaging capabilities

Abstract

Temporal focusing two-photon microscopy enables high resolution imaging of fine structures in vivo over a large volume. A limitation of temporal focusing is that signal-to-background ratio and resolution degrade rapidly with increasing imaging depth. This degradation originates from the scattered emission photons are widely distributed resulting in a strong background. We have developed Multiline Orthogonal Scanning Temporal Focusing (mosTF) microscopy that overcomes this problem. mosTF captures a sequence of images at each scan location of the excitation line, followed by a reconstruction algorithm reassigns scattered photons back to the correct scan position. We demonstrate mosTF by acquiring mice neuronal images in vivo. Our results show remarkably improvements with mosTF for in vivo brain imaging while maintaining its speed advantage.