Direct optical nanoscopy with axially localized detection

TL;DR

This paper introduces DONALD, a 3D nanoscopy technique combining dSTORM with supercritical angle fluorescence detection, achieving 20 nm axial localization precision for super-resolution imaging.

Contribution

It presents a novel method that exploits evanescent light in emission to enhance 3D localization accuracy in fluorescence microscopy.

Findings

Achieves 20 nm isotropic 3D localization precision.

Combines dSTORM with SAF detection for improved axial resolution.

Demonstrates effective 3D super-resolution imaging.

Abstract

Evanescent light excitation is widely used in super-resolution fluorescence microscopy to confine light and reduce background noise. Herein we propose a method of exploiting evanescent light in the context of emission. When a fluorophore is located in close proximity to a medium with a higher refractive index, its near-field component is converted into light that propagates beyond the critical angle. This so-called Supercritical Angle Fluorescence (SAF) can be captured using a hig-NA objective and used to determine the axial position of the fluorophore with nanometer precision. We introduce a new technique for 3D nanoscopy that combines direct STochastic Optical Reconstruction Microscopy (dSTORM) imaging with dedicated detection of SAF emission. We demonstrate that our approach of a Direct Optical Nanoscopy with Axially Localized Detection (DONALD) yields a typical isotropic 3D localization precision of 20 nm.