Super-resolution deep imaging with hollow Bessel beam STED microscopy

TL;DR

This paper introduces a deep imaging STED microscopy technique using a hollow Bessel beam for depletion, achieving significantly greater imaging depths while maintaining super-resolution, surpassing standard STED limitations.

Contribution

The study presents a novel GB-STED method that enhances imaging depth and resolution in thick specimens, addressing aberration and scattering challenges in deep cellular imaging.

Findings

Imaging depth increased to ~155μm in agarose

Consistent super-resolution at greater depths

Outperforms standard STED in deep tissue imaging

Abstract

Stimulated emission depletion (STED) microscopy has become a powerful imaging and localized excitation method beating the diffraction barrier for improved lateral spatial resolution in cellular imaging, lithography, etc. Due to specimen-induced aberrations and scattering distortion, it has been a great challenge for STED to maintain consistent lateral resolution deeply inside the specimens. Here we report on a deep imaging STED microscopy by using Gaussian beam for excitation and hollow Bessel beam for depletion (GB-STED). The proposed scheme shows the improved imaging depth up to ~155{\mu}m in solid agarose sample, ~115{\mu}m in PDMS and ~100{\mu}m in phantom of gray matter in brain tissue with consistent super resolution, while the standard STED microscopy shown a significantly reduced lateral resolution at the same imaging depth. The results indicate the excellent imaging penetration capability of GB-STED, making it a promising tool for deep 3D imaging optical nanoscopy and laser fabrication.