Fast volumetric imaging with patterned illumination via digital micro-mirror device-based temporal focusing...

Area of Science:

• Biophysics
• Optical Microscopy
• 3D Imaging

Background:

• Temporal focusing multiphoton microscopy (TFMPM) offers precise axial confinement for fast 3D imaging.
• Existing methods may lack the speed required for dynamic biological processes.

Purpose of the Study:

• To develop and demonstrate a fast volumetric imaging technique using digital micromirror device (DMD)-based TFMPM.
• To enable real-time 3D observation of dynamic microscopic phenomena.

Main Methods:

• Synchronized system integrating a DMD-based TFMPM with an electron multiplying charge-coupled device (EMCCD) and a piezoelectric stage.
• High-speed axial scanning for volumetric data acquisition.
• Dynamic patterned illumination for noise reduction.

Main Results:

• Achieved a volumetric imaging rate of 30 volumes per second, leveraging an EMCCD frame rate exceeding 400 fps.
• Successfully observed the 3D Brownian motion of one-micron fluorescent beads in real-time.
• Demonstrated background noise rejection using dynamic HiLo structural multiphoton microscopy with high-speed DMD illumination.

Conclusions:

• The developed DMD-based TFMPM system significantly enhances volumetric imaging speed for 3D multiphoton microscopy.
• This technique provides a powerful tool for studying fast dynamic processes at the microscale.
• The integration of high-speed illumination and detection enables improved image quality and noise suppression.