Self-Driving Microscopes: AI Meets Super-Resolution Microscopy.

The integration of Machine Learning (ML) with super-resolution microscopy represents a transformative advancement in biomedical research. Recent advances in ML, particularly deep learning (DL), have significantly enhanced image processing tasks, such as denoising and reconstruction. This review explores the growing potential of automation in super-resolution microscopy, focusing on how DL can enable autonomous imaging tasks.

A High-Resolution Microscopy System for Biological Studies of Cold-Adapted Species Under Physiological Conditions.

The Antarctic seabed harbors significant biodiversity, and almost 90% of oceanic environments are permanently below 5 °C (i.e., deep sea and polar regions).

Enzyme-Responsive DNA Condensates.

Membrane-less compartments and organelles are widely acknowledged for their role in regulating cellular processes, and there is an urgent need to harness their full potential as both structural and functional elements of synthetic cells. Despite rapid progress, synthetically recapitulating the nonequilibrium, spatially distributed responses of natural membrane-less organelles remains elusive. Here, we demonstrate that the activity of nucleic-acid cleaving enzymes can be localized within DNA-based membrane-less compartments by sequestering the respective DNA or RNA substrates.

Connexin 30 locally controls actin cytoskeleton and mechanical remodeling in motile astrocytes.

During brain maturation, astrocytes establish complex morphologies unveiling intense structural plasticity. Connexin 30 (Cx30), a gap-junction channel-forming protein expressed postnatally, dynamically regulates during development astrocyte morphological properties by controlling ramification and extension of fine processes. However, the underlying mechanisms remain unexplored.

Mutation of the ALS-/FTD-Associated RNA-Binding Protein FUS Affects Axonal Development.

Aberrant condensation and localization of the RNA-binding protein (RBP) fused in sarcoma (FUS) occur in variants of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Changes in RBP function are commonly associated with changes in axonal cytoskeletal organization and branching in neurodevelopmental disorders. Here, we asked whether branching defects also occur in vivo in a model of FUS-associated disease.

Fast, multicolour optical sectioning over extended fields of view with patterned illumination and 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. However, this optical sectioning technique is currently limited by image reconstruction artefacts and poor performance at low signal-to-noise ratios. We combine multicolour interferometric pattern generation with machine learning to achieve high-contrast, real-time reconstruction of image data that is robust to background noise and sample motion.

Molecular Mechanisms of Cationic Fusogenic Liposome Interactions with Bacterial Envelopes.

Although fusogenic liposomes offer a promising approach for the delivery of antibiotic payloads across the cell envelope of Gram-negative bacteria, there is still a limited understanding of the individual nanocarrier interactions with the bacterial target. Using super-resolution microscopy, we characterize the interaction dynamics of positively charged fusogenic liposomes with Gram-negative () and Gram-positive () bacteria. The liposomes merge with the outer membrane (OM) of Gram-negative bacteria, while attachment or lipid internalization is observed in Gram-positive cells.

Diversity of dynamic voltage patterns in neuronal dendrites revealed by nanopipette electrophysiology.

Dendrites and dendritic spines are the essential cellular compartments in neuronal communication, conveying information through transient voltage signals. Our understanding of these compartmentalized voltage dynamics in fine, distal neuronal dendrites remains poor due to the difficulties inherent to accessing and stably recording from such small, nanoscale cellular compartments for a sustained time. To overcome these challenges, we use nanopipettes that permit long and stable recordings directly from fine neuronal dendrites.

α-Synuclein fibril and synaptic vesicle interactions lead to vesicle destruction and increased lipid-associated fibril uptake into iPSC-derived neurons.

Monomeric alpha-synuclein (aSyn) is a well characterised protein that importantly binds to lipids. aSyn monomers assemble into amyloid fibrils which are localised to lipids and organelles in insoluble structures found in Parkinson's disease patient's brains. Previous work to address pathological aSyn-lipid interactions has focused on using synthetic lipid membranes, which lack the complexity of physiological lipid membranes.

Multiparametric Sensing of Outer Membrane Vesicle-Derived Supported Lipid Bilayers Demonstrates the Specificity of Bacteriophage Interactions.

The use of bacteriophages, viruses that specifically infect bacteria, as antibiotics has become an area of great interest in recent years as the effectiveness of conventional antibiotics recedes. The detection of phage interactions with specific bacteria in a rapid and quantitative way is key for identifying phages of interest for novel antimicrobials. Outer membrane vesicles (OMVs) derived from Gram-negative bacteria can be used to make supported lipid bilayers (SLBs) and therefore membrane models that contain naturally occurring components of the bacterial outer membrane.

ERnet: a tool for the semantic segmentation and quantitative analysis of endoplasmic reticulum topology.

The ability to quantify structural changes of the endoplasmic reticulum (ER) is crucial for understanding the structure and function of this organelle. However, the rapid movement and complex topology of ER networks make this challenging. Here, we construct a state-of-the-art semantic segmentation method that we call ERnet for the automatic classification of sheet and tubular ER domains inside individual cells.

Feasibility of Coacervate-Like Nanostructure for Instant Drug Nanoformulation.

Despite the enormous advancements in nanomedicine research, a limited number of nanoformulations are available on the market, and few have been translated to clinics. An easily scalable, sustainable, and cost-effective manufacturing strategy and long-term stability for storage are crucial for successful translation. Here, we report a system and method to instantly formulate NF achieved with a nanoscale polyelectrolyte coacervate-like system, consisting of anionic pseudopeptide poly(l-lysine isophthalamide) derivatives, polyethylenimine, and doxorubicin (Dox) via simple "mix-and-go" addition of precursor solutions in seconds.

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 the real space coordinate frame in which the sample is moved. This makes live viewing and practical operation of such microscopes difficult.

Biosensor for Multimodal Characterization of an Essential ABC Transporter for Next-Generation Antibiotic Research.

As the threat of antibiotic resistance increases, there is a particular focus on developing antimicrobials against pathogenic bacteria whose multidrug resistance is especially entrenched and concerning. One such target for novel antimicrobials is the ATP-binding cassette (ABC) transporter MsbA that is present in the plasma membrane of Gram-negative pathogenic bacteria where it is fundamental to the survival of these bacteria. Supported lipid bilayers (SLBs) are useful in monitoring membrane protein structure and function since they can be integrated with a variety of optical, biochemical, and electrochemical techniques.

Single-Molecule Sizing through Nanocavity Confinement.

An approach relying on nanocavity confinement is developed in this paper for the sizing of nanoscale particles and single biomolecules in solution. The approach, termed nanocavity diffusional sizing (NDS), measures particle residence times within nanofluidic cavities to determine their hydrodynamic radii. Using theoretical modeling and simulations, we show that the residence time of particles within nanocavities above a critical time scale depends on the diffusion coefficient of the particle, which allows the estimation of the particle's size.

Machine learning assisted interferometric structured illumination microscopy for dynamic biological imaging.

Structured Illumination Microscopy, SIM, is one of the most powerful optical imaging methods available to visualize biological environments at subcellular resolution. Its limitations stem from a difficulty of imaging in multiple color channels at once, which reduces imaging speed. Furthermore, there is substantial experimental complexity in setting up SIM systems, preventing a widespread adoption.

Near-native state imaging by cryo-soft-X-ray tomography reveals remodelling of multiple cellular organelles during HSV-1 infection.

Herpes simplex virus-1 (HSV-1) is a large, enveloped DNA virus and its assembly in the cell is a complex multi-step process during which viral particles interact with numerous cellular compartments such as the nucleus and organelles of the secretory pathway. Transmission electron microscopy and fluorescence microscopy are commonly used to study HSV-1 infection. However, 2D imaging limits our understanding of the 3D geometric changes to cellular compartments that accompany infection and sample processing can introduce morphological artefacts that complicate interpretation.

Protein Condensation, Cellular Organization, and Spatiotemporal Regulation of Cytoplasmic Properties.

The cytoplasm is an aqueous, highly crowded solution of active macromolecules. Its properties influence the behavior of proteins, including their folding, motion, and interactions. In particular, proteins in the cytoplasm can interact to form phase-separated assemblies, so-called biomolecular condensates.

Fluorescent Nanoparticles for Super-Resolution Imaging.

Super-resolution imaging techniques that overcome the diffraction limit of light have gained wide popularity for visualizing cellular structures with nanometric resolution. Following the pace of hardware developments, the availability of new fluorescent probes with superior properties is becoming ever more important. In this context, fluorescent nanoparticles (NPs) have attracted increasing attention as bright and photostable probes that address many shortcomings of traditional fluorescent probes.

Nanoscale Features of Tunable Bacterial Outer Membrane Models Revealed by Correlative Microscopy.

The rise of antibiotic resistance is a growing worldwide human health issue, with major socioeconomic implications. An understanding of the interactions occurring at the bacterial membrane is crucial for the generation of new antibiotics. Supported lipid bilayers (SLBs) made from reconstituted lipid vesicles have been used to mimic these membranes, but their utility has been restricted by the simplistic nature of these systems.

A method for the fast and photon-efficient analysis of time-domain fluorescence lifetime image data over large dynamic ranges.

Fluorescence lifetime imaging (FLIM) allows the quantification of sub-cellular processes in situ, in living cells. A number of approaches have been developed to extract the lifetime from time-domain FLIM data, but they are often limited in terms of speed, photon efficiency, precision or the dynamic range of lifetimes they can measure. Here, we focus on one of the best performing methods in the field, the centre-of-mass method (CMM), that conveys advantages in terms of speed and photon efficiency over others.

Intracellular Aβ42 Aggregation Leads to Cellular Thermogenesis.

The aggregation of Aβ42 is a hallmark of Alzheimer's disease. It is still not known what the biochemical changes are inside a cell which will eventually lead to Aβ42 aggregation. Thermogenesis has been associated with cellular stress, the latter of which may promote aggregation.

Tuning riboflavin derivatives for photodynamic inactivation of pathogens.

The development of effective pathogen reduction strategies is required due to the rise in antibiotic-resistant bacteria and zoonotic viral pandemics. Photodynamic inactivation (PDI) of bacteria and viruses is a potent reduction strategy that bypasses typical resistance mechanisms. Naturally occurring riboflavin has been widely used in PDI applications due to efficient light-induced reactive oxygen species (ROS) release.