Compressive three-dimensional super-resolution microscopy with speckle-saturated fluorescence excitation

TL;DR

This paper introduces a novel 3D super-resolution microscopy technique using speckle-saturated fluorescence excitation combined with compressed sensing, enabling faster imaging with less photobleaching.

Contribution

It presents the first 3D super-resolution nSIM method leveraging speckle patterns and compressed sensing for efficient, high-contrast imaging of biological samples.

Findings

Achieved 3D super-resolution imaging with single 2D scan

Reduced photobleaching and acquisition time

Utilized speckle patterns for contrast and orthogonality

Abstract

Nonlinear structured illumination microscopy (nSIM) is an effective approach for super-resolution wide-field fluorescence microscopy with a theoretically unlimited resolution. In nSIM, carefully designed, highly-contrasted illumination patterns are combined with the saturation of an optical transition to enable sub-diffraction imaging. While the technique proved useful for two-dimensional imaging, extending it to three-dimensions (3D) is challenging due to the fading/fatigue of organic fluorophores under intense cycling conditions. Here, we present a compressed sensing approach that allows for the first time 3D sub-diffraction nSIM of cultured cells by saturating fluorescence excitation. Exploiting the natural orthogonality of transverse speckle illumination planes, 3D probing of the sample is achieved by a single two-dimensional scan. Fluorescence contrast under saturated excitation is ensured by the inherent high density of intensity minima associated with optical vortices in polarized speckle patterns. Compressed speckle microscopy is thus a simple approach that enables 3D super-resolved nSIM imaging with potentially considerably reduced acquisition time and photobleaching.les fast 3D super-resolved imaging with considerably minimized photo-bleaching.