Rapid 3D image scanning microscopy with multi-spot excitation and double-helix point spread function detection

Area of Science:

• Microscopy and Imaging Technologies
• Optical Physics
• Biomedical Engineering

Background:

• Refocusing after Scanning using Helical phase engineering (RESCH) microscopy offers volumetric data from single 2D scans.
• Practical RESCH applications are hindered by slow acquisition speeds and limited spatial resolution.

Purpose of the Study:

• To enhance image acquisition speed and spatial resolution by combining RESCH with multifocal structured illumination microscopy (MSIM).
• To develop a novel 3D image scanning microscopy technique for improved volumetric imaging.

Main Methods:

• Integration of a double-helix point spread function (DH-PSF) using a phase mask for volumetric data.
• Generation of sparse multifocal illumination patterns via a digital micromirror device (DMD) for parallel 3D imaging.
• Application of Richardson-Lucy deconvolution and pixel reassignment for enhanced spatial resolution and depth estimation.

Main Results:

• The combined RESCH-MSIM system achieves faster data acquisition and a larger field of view compared to conventional RESCH.
• Demonstrated significant improvements in spatial resolution for both 3D specimen imaging and depth estimation.
• Successfully recorded comprehensive 3D specimen and depth information from a single multi-spot 2D planar scan.

Conclusions:

• The proposed 3D image scanning microscopy effectively overcomes the limitations of RESCH.
• This novel approach provides a powerful tool for high-speed, high-resolution 3D imaging in various scientific applications.
• Experimental validation confirms the system's capability for advanced volumetric imaging.