Long axial-range double-helix point spread functions for 3D volumetric super-resolution imaging.

Area of Science:

• Biophysics
• Optical Microscopy
• Nanotechnology

Background:

• Single-molecule localization microscopy (SMLM) achieves super-resolution imaging beyond light diffraction limits.
• Engineered point spread functions (PSFs) extend SMLM to 3D imaging, but challenges remain for thick samples like mammalian cells.

Purpose of the Study:

• To simplify 3D super-resolution imaging workflows in thick biological samples.
• To demonstrate the utility of long axial-range double-helix (DH)-PSFs for stitching-free volumetric imaging.

Main Methods:

• Experimental benchmarking of DH-PSF localization precision using fluorescent beads.
• 3D SMLM of mammalian cells (U-2 OS) using DNA-PAINT to image lamin B1.
• Application of a deep learning algorithm for localizing dense emitters.

Main Results:

• DH-PSFs were characterized for precision, resolution, and imaging speed.
• Successful 3D super-resolution imaging of the nuclear lamina protein lamin B1 in U-2 OS cells.
• Deep learning significantly enhanced imaging speed and resolution for dense emitters.

Conclusions:

• Long axial-range DH-PSFs enable stitching-free 3D super-resolution imaging of entire mammalian cells.
• The developed method simplifies experimental and analysis procedures for nanoscale structural information.
• This approach facilitates volumetric nanoscale imaging of complex biological structures.