Mechanically sheared axially swept light-sheet microscopy.

We present a mechanically sheared image acquisition format for upright and open-top light-sheet microscopes that automatically places data in its proper spatial context. This approach, which reduces computational post-processing and eliminates unnecessary interpolation or duplication of the data, is demonstrated on an upright variant of axially swept light-sheet microscopy (ASLM) that achieves a field of view, measuring 774 × 435 microns, that is 3.2-fold larger than previous models and a raw and isotropic resolution of ∼460 nm.

Mechanically Sheared Axially Swept Light-Sheet Microscopy.

We present a mechanically sheared image acquisition format for upright and open-top light-sheet microscopes that automatically places data in its proper spatial context. This approach, which reduces computational post-processing and eliminates unnecessary interpolation or duplication of the data, is demonstrated on an upright variant of Axially Swept Light-Sheet Microscopy (ASLM) that achieves a field of view, measuring 774 x 435 microns, that is 3.2-fold larger than previous models and a raw and isotropic resolution of ∼420 nm.

navigate: an open-source platform for smart light-sheet microscopy.

is a turnkey, open-source software solution designed to enhance light-sheet fluorescence microscopy (LSFM) by integrating smart microscopy techniques into a user-friendly framework. It enables automated, intelligent imaging with a Python-based control system that supports GUI-reconfigurable acquisition routines and the integration of diverse hardware sets. As a comprehensive package, navigate democratizes access to advanced LSFM capabilities, facilitating the development and implementation of smart microscopy workflows without requiring deep programming knowledge or specialized expertise in light-sheet microscopy.

Correlative Conventional and Super-resolution Photoactivated Localization Microscopy (PALM) Imaging to Characterize Chromatin Structure and Dynamics in Live Mammalian Cells.

A fundamental understanding of gene regulation requires a quantitative characterization of the spatial organization and dynamics of chromatin. The advent of fluorescence super-resolution microscopy techniques such as photoactivated localization microscopy (PALM) presented a breakthrough to visualize structural features with a resolution of ~20 nm in fixed cells. However, until recently the long acquisition time of super-resolution images prevented high-resolution measurements in living cells due to spreading of localizations caused by chromatin motion.

Organelle interactions compartmentalize hepatic fatty acid trafficking and metabolism.

Organelle interactions play a significant role in compartmentalizing metabolism and signaling. Lipid droplets (LDs) interact with numerous organelles, including mitochondria, which is largely assumed to facilitate lipid transfer and catabolism. However, quantitative proteomics of hepatic peridroplet mitochondria (PDM) and cytosolic mitochondria (CM) reveals that CM are enriched in proteins comprising various oxidative metabolism pathways, whereas PDM are enriched in proteins involved in lipid anabolism.

Efficient Cross-Correlation Filtering of One- and Two-Color Single Molecule Localization Microscopy Data.

Single molecule localization microscopy has become a prominent technique to quantitatively study biological processes below the optical diffraction limit. By fitting the intensity profile of single sparsely activated fluorophores, which are often attached to a specific biomolecule within a cell, the locations of all imaged fluorophores are obtained with ∼20 nm resolution in the form of a coordinate table. While rendered super-resolution images reveal structural features of intracellular structures below the optical diffraction limit, the ability to further analyze the molecular coordinates presents opportunities to gain additional quantitative insights into the spatial distribution of a biomolecule of interest.

PS F: Polarized Spiral Point Spread Function for Single-Shot 3D Sensing.

We propose a compact snapshot monocular depth estimation technique that relies on an engineered point spread function (PSF). Traditional approaches used in microscopic super-resolution imaging such as the Double-Helix PSF (DHPSF) are ill-suited for scenes that are more complex than a sparse set of point light sources. We show, using the Cramér-Rao lower bound, that separating the two lobes of the DHPSF and thereby capturing two separate images leads to a dramatic increase in depth accuracy.

Characterizing locus specific chromatin structure and dynamics with correlative conventional and super-resolution imaging in living cells.

The dynamic rearrangement of chromatin is critical for gene regulation, but mapping both the spatial organization of chromatin and its dynamics remains a challenge. Many structural conformations are too small to be resolved via conventional fluorescence microscopy and the long acquisition time of super-resolution photoactivated localization microscopy (PALM) precludes the structural characterization of chromatin below the optical diffraction limit in living cells due to chromatin motion. Here we develop a correlative conventional fluorescence and PALM imaging approach to quantitatively map time-averaged chromatin structure and dynamics below the optical diffraction limit in living cells.

The Role of Protein Loss and Denaturation in Determining Outcomes of Heating, Cryotherapy, and Irreversible Electroporation on Cardiomyocytes.

Atrial fibrillation (AF) currently affects millions of people in the U.S. alone.

Spectroscopic and calorimetric evaluation of chemically induced protein denaturation in HuH-7 liver cancer cells and impact on cell survival.

Solid tumors such as hepatocellular carcinoma are very often not amenable to chemotherapy and radiotherapy. Local ablation methods, including chemical ablation with absolute ethanol, are therefore an option for treatment but lack of information about the mechanism of devitalization leading to cell death is a hindrance to further adoption. Systemic toxicity also has limited the amount of ethanol that can be used in a single treatment session.