Giroussens, G. | Search bioRxiv

bioRxiv2025

Super-resolved live imaging of thick biological samples with 3D Random Illumination Microscopy (3D-RIM)

Super-resolved volume imaging of thick, live specimens is greatly hampered by sample induced aberrations and out-of-focus fluorescence. In this work, using a combination of speckled illuminations, remote focusing, three-dimensional photon reassignement and variance processing, we obtained volume images with super-resolution (110 nm transverse, 270 nm axial) on thin samples and maintained a high contrast and high resolution level throughout tens of microns of highly aberrant tissues and up to hundreds of microns in collagen scaffolds.

bioengineering↗

bioRxiv2023

Extended-depth of field random illumination microscopy, EDF-RIM, provides super-resolved projective imaging

The ultimate aim of fluorescence microscopy is to achieve high-resolution imaging of increasingly larger biological samples. Extended depth of field presents a potential solution to accelerate imaging of large samples when compression of information along the optical axis is not detrimental to the interpretation of images. We have implemented an Extended Depth of Field (EDF) approach in a Random Illumination Microscope (RIM). RIM uses multiple speckled illuminations and variance data processing to double the resolution. It is particularly adapted to the imaging of thick samples as it does not require the knowledge of illumination patterns. We demonstrate highly-resolved projective images of biological tissues and cells. Compared to a sequential scan of the imaged volume with conventional 2D-RIM, EDF-RIM allows an order of magnitude improvement in speed and light dose reduction, with comparable resolution. As the axial information is lost in an EDF modality, we propose a method to retrieve the sample topography for samples that are organized in cell sheets.

bioengineering↗