MicroMagnify: A Multiplexed Expansion Microscopy Method for Pathogens and Infected Tissues.

Super-resolution optical imaging tools are crucial in microbiology to understand the complex structures and behavior of microorganisms such as bacteria, fungi, and viruses. However, the capabilities of these tools, particularly when it comes to imaging pathogens and infected tissues, remain limited. MicroMagnify (µMagnify) is developed, a nanoscale multiplexed imaging method for pathogens and infected tissues that are derived from an expansion microscopy technique with a universal biomolecular anchor. The combination of heat denaturation and enzyme cocktails essential is found for robust cell wall digestion and expansion of microbial cells and infected tissues without distortion. µMagnify efficiently retains biomolecules suitable for high-plex fluorescence imaging with nanoscale precision. It demonstrates up to eightfold expansion with µMagnify on a broad range of pathogen-containing specimens, including bacterial and fungal biofilms, infected culture cells, fungus-infected mouse tone, and formalin-fixed paraffin-embedded human cornea infected by various pathogens. Additionally, an associated virtual reality tool is developed to facilitate the visualization and navigation of complex 3D images generated by this method in an immersive environment allowing collaborative exploration among researchers worldwide. µMagnify is a valuable imaging platform for studying how microbes interact with their host systems and enables the development of new diagnosis strategies against infectious diseases.

MicroMagnify: a multiplexed expansion microscopy method for pathogens and infected tissues.

Super-resolution optical imaging tools are crucial in microbiology to understand the complex structures and behavior of microorganisms such as bacteria, fungi, and viruses. However, the capabilities of these tools, particularly when it comes to imaging pathogens and infected tissues, remain limited. We developed µMagnify, a nanoscale multiplexed imaging method for pathogens and infected tissues that are derived from an expansion microscopy technique with a universal biomolecular anchor. We formulated an enzyme cocktail specifically designed for robust cell wall digestion and expansion of microbial cells without distortion while efficiently retaining biomolecules suitable for high-plex fluorescence imaging with nanoscale precision. Additionally, we developed an associated virtual reality tool to facilitate the visualization and navigation of complex three-dimensional images generated by this method in an immersive environment allowing collaborative exploration among researchers around the world. µMagnify is a valuable imaging platform for studying how microbes interact with their host systems and enables development of new diagnosis strategies against infectious diseases.

Magnify is a universal molecular anchoring strategy for expansion microscopy.

Expansion microscopy enables nanoimaging with conventional microscopes by physically and isotropically magnifying preserved biological specimens embedded in a crosslinked water-swellable hydrogel. Current expansion microscopy protocols require prior treatment with reactive anchoring chemicals to link specific labels and biomolecule classes to the gel. We describe a strategy called Magnify, which uses a mechanically sturdy gel that retains nucleic acids, proteins and lipids without the need for a separate anchoring step. Magnify expands biological specimens up to 11 times and facilitates imaging of cells and tissues with effectively around 25-nm resolution using a diffraction-limited objective lens of about 280 nm on conventional optical microscopes or with around 15 nm effective resolution if combined with super-resolution optical fluctuation imaging. We demonstrate Magnify on a broad range of biological specimens, providing insight into nanoscopic subcellular structures, including synaptic proteins from mouse brain, podocyte foot processes in formalin-fixed paraffin-embedded human kidney and defects in cilia and basal bodies in drug-treated human lung organoids.

Repurposing Peptide Nanomaterials as Synthetic Biomolecular Condensates in Bacteria.

Peptide nanomaterials exhibit diverse applications in vitro, such as drug delivery. Here, we consider the utility of de novo peptide nanomaterials to organize biochemistry within the bacterial cytoplasm. Toward this goal, we discovered that ABC coiled-coil triblock peptides form gel-like biomolecular condensates with a csat of 10 µM in addition to their well-known hydrogel-forming capabilities. Expression of the coiled-coil triblock peptides in bacteria leads to cell pole accumulation via a nucleoid occlusion mechanism. We then provide a proof of principle that these synthetic biomolecular condensates could sequester clients at the cell pole. Finally, we demonstrate that triblock peptides and another biomolecular condensate, RNase E, phase-separate as distinct protein-rich assemblies in vitro and in vivo. These results reveal the potential of using peptide nanomaterials to divide the bacterial cytoplasm into distinct subcellular zones with future metabolic engineering and synthetic biology applications.

The Role of Extracellular Vesicles in the Pathogenesis and Treatment of Autoimmune Disorders.

Extracellular vesicles (EVs) are important players in autoimmune diseases, both in disease pathogenesis and as potential treatments. EVs can transport autoimmune triggers throughout the body, facilitating the process of antigen presentation. Understanding the link between cellular stress and EV biogenesis and intercellular trafficking will advance our understanding of autoimmune diseases. In addition, EVs can also be effective treatments for autoimmune diseases. The diversity of cell types that produce EVs leads to a wide range of molecules to be present in EVs, and thus EVs have a wide range of physiological effects. EVs derived from dendritic cells or mesenchymal stem cells have been shown to reduce inflammation. Since many autoimmune treatments are focused only on symptom management, EVs present a promising avenue for potential treatments. This review looks at the different roles EVs can play in autoimmune diseases, from disease pathology to diagnosis and treatment. We also overview various methodologies in isolating or generating EVs and look to the future for possible applications of EVs in autoimmune diseases.