Combined Multi-Plane Tomographic Phase Retrieval and Stochastic Optical Fluctuation Imaging for 4D Cell Microscopy

TL;DR

This paper introduces a novel 4D microscopy platform combining multi-plane phase tomography and fluorescence imaging, enabling high-speed, high-resolution live cell imaging without sacrificing temporal or spatial detail.

Contribution

The work presents a non-iterative, multi-plane phase reconstruction method integrated with fluorescence super-resolution imaging in a single, high-speed microscope platform.

Findings

Achieved 3D live cell imaging at 200 Hz.

Integrated phase tomography with fluorescence super-resolution.

Enabled simultaneous high-speed, high-resolution 4D imaging.

Abstract

Super-resolution fluorescence microscopy provides unprecedented insight into cellular and subcellular structures. However, going "beyond the diffraction barrier" comes at a price since most far-field super-resolution imaging techniques trade temporal for spatial super-resolution. We propose the combination of a novel label-free white light quantitative phase tomography with fluorescence imaging to provide high-speed imaging and spatial super-resolution. The non-iterative phase reconstruction relies on the acquisition of single images at each z-location and thus enables straightforward 3D phase imaging using a classical microscope. We realized multi-plane imaging using a customized prism for the simultaneous acquisition of 8 planes. This allowed us to not only image live cells in 3D at up to 200 Hz, but also to integrate fluorescence super-resolution optical fluctuation imaging within the same optical instrument. This 4D microscope platform unifies the sensitivity and high temporal resolution of phase tomography with the specificity and high spatial resolution of fluorescence imaging.