Three-dimensional in vivo scanning microscopy with inertia-free focus control

Area of Science:

• Biomedical optics
• Microscopy
• Neuroscience

Background:

• High-resolution 3D imaging is crucial for studying biological tissue morphology and function.
• Conventional microscopy methods often face limitations with moving objectives, affecting image quality and speed.

Purpose of the Study:

• To present a novel laser scanning two-photon microscope with remote, motionless focus control.
• To enable inertia-free 3D imaging with precise control over the focal plane.

Main Methods:

• Utilized a spatial light modulator to shape the laser wavefront, controlling the excitation spot's position.
• Implemented remote and motionless focus control for z-axis movement.
• Applied the system for in vivo 3D imaging of fluorescent cells in the mouse neocortex.

Main Results:

• Achieved z movements of tens to hundreds of micrometers with minimal changes to the point spread function.
• Demonstrated high-resolution, inertia-free 3D imaging in biological samples.
• Successfully imaged fluorescent cells in the mouse neocortex in vivo.

Conclusions:

• The developed two-photon microscope overcomes limitations of objective-moving systems.
• This technique offers a powerful tool for advanced morphological and functional investigations of biological tissues.
• Enables high-quality, inertia-free 3D imaging for in vivo neuroscience research.