Fast, multicolour optical sectioning over extended fields of view by combining interferometric SIM with machine learning

Structured illumination can reject out-of-focus signal from a sample, enabling high-speed and high-contrast imaging over large areas with widefield detection optics. Currently, this optical-sectioning technique is limited by image reconstruction artefacts and the need for sequential imaging of multiple colour channels. We combine multicolour interferometric pattern generation with machine-learning processing, permitting high-contrast, real-time reconstruction of image data. The method is insensitive to background noise and unevenly phase-stepped illumination patterns. We validate the method in silico and demonstrate its application on diverse specimens, ranging from fixed and live biological cells to synthetic biosystems, imaging at up to 37 Hz across a 44 x 44 $\mu m^2$ field of view.

An open-source software package for on-the-fly deskewing and live viewing of volumetric lightsheet microscopy data

Oblique plane microscopy, OPM, is a form of lightsheet microscopy that permits volumetric imaging of biological samples at high temporal and spatial resolution. However, the imaging geometry of OPM, and related variants of light sheet microscopy, distorts the coordinate frame of the presented image sections with respect to real space coordinate frame in which the sample is moved to navigate to regions of interest. This makes live viewing and practical operation of such microscopes difficult. We present an open-source software package that utilises GPU acceleration and multiprocessing to transform the display of OPM imaging data in real time to produce live views that mimic that produced by standard widefield microscopes. Image stacks can be acquired, processed and plotted at rates of several Hz, making live operation of OPMs, and similar microscopes, more user friendly and intuitive.

Spatio-temporal Vision Transformer for Super-resolution Microscopy

Structured illumination microscopy (SIM) is an optical super-resolution technique that enables live-cell imaging beyond the diffraction limit. Reconstruction of SIM data is prone to artefacts, which becomes problematic when imaging highly dynamic samples because previous methods rely on the assumption that samples are static. We propose a new transformer-based reconstruction method, VSR-SIM, that uses shifted 3-dimensional window multi-head attention in addition to channel attention mechanism to tackle the problem of video super-resolution (VSR) in SIM. The attention mechanisms are found to capture motion in sequences without the need for common motion estimation techniques such as optical flow. We take an approach to training the network that relies solely on simulated data using videos of natural scenery with a model for SIM image formation. We demonstrate a use case enabled by VSR-SIM referred to as rolling SIM imaging, which increases temporal resolution in SIM by a factor of 9. Our method can be applied to any SIM setup enabling precise recordings of dynamic processes in biomedical research with high temporal resolution.

ML-SIM: A deep neural network for reconstruction of structured illumination microscopy images

Structured illumination microscopy (SIM) has become an important technique for optical super-resolution imaging because it allows a doubling of image resolution at speeds compatible for live-cell imaging. However, the reconstruction of SIM images is often slow and prone to artefacts. Here we propose a versatile reconstruction method, ML-SIM, which makes use of machine learning. The model is an end-to-end deep residual neural network that is trained on a simulated data set to be free of common SIM artefacts. ML-SIM is thus robust to noise and irregularities in the illumination patterns of the raw SIM input frames. The reconstruction method is widely applicable and does not require the acquisition of experimental training data. Since the training data are generated from simulations of the SIM process on images from generic libraries the method can be efficiently adapted to specific experimental SIM implementations. The reconstruction quality enabled by our method is compared with traditional SIM reconstruction methods, and we demonstrate advantages in terms of noise, reconstruction fidelity and contrast for both simulated and experimental inputs. In addition, reconstruction of one SIM frame typically only takes ~100ms to perform on PCs with modern Nvidia graphics cards, making the technique compatible with real-time imaging. The full implementation and the trained networks are available at http://ML-SIM.com.