Hasil untuk "Microscopy"

Claudia Errico, J. Pierre, S. Pezet et al.

Keiichi Tanaka, K. Maeda

J. Grimm, Brian P. English, Jiji Chen et al.

Specific labeling of biomolecules with bright fluorophores is the keystone of fluorescence microscopy. Genetically encoded self-labeling tag proteins can be coupled to synthetic dyes inside living cells, resulting in brighter reporters than fluorescent proteins. Intracellular labeling using these techniques requires cell-permeable fluorescent ligands, however, limiting utility to a small number of classic fluorophores. Here we describe a simple structural modification that improves the brightness and photostability of dyes while preserving spectral properties and cell permeability. Inspired by molecular modeling, we replaced the N,N-dimethylamino substituents in tetramethylrhodamine with four-membered azetidine rings. This addition of two carbon atoms doubles the quantum efficiency and improves the photon yield of the dye in applications ranging from in vitro single-molecule measurements to super-resolution imaging. The novel substitution is generalizable, yielding a palette of chemical dyes with improved quantum efficiencies that spans the UV and visible range.

J. Frank, M. Radermacher, M. Radermacher et al.

C. Suloway, J. Pulokas, D. Fellmann et al.

R. Power, J. Huisken

D. Sage, Laurène Donati, F. Soulez et al.

Images in fluorescence microscopy are inherently blurred due to the limit of diffraction of light. The purpose of deconvolution microscopy is to compensate numerically for this degradation. Deconvolution is widely used to restore fine details of 3D biological samples. Unfortunately, dealing with deconvolution tools is not straightforward. Among others, end users have to select the appropriate algorithm, calibration and parametrization, while potentially facing demanding computational tasks. To make deconvolution more accessible, we have developed a practical platform for deconvolution microscopy called DeconvolutionLab. Freely distributed, DeconvolutionLab hosts standard algorithms for 3D microscopy deconvolution and drives them through a user-oriented interface. In this paper, we take advantage of the release of DeconvolutionLab2 to provide a complete description of the software package and its built-in deconvolution algorithms. We examine several standard algorithms used in deconvolution microscopy, notably: Regularized inverse filter, Tikhonov regularization, Landweber, Tikhonov-Miller, Richardson-Lucy, and fast iterative shrinkage-thresholding. We evaluate these methods over large 3D microscopy images using simulated datasets and real experimental images. We distinguish the algorithms in terms of image quality, performance, usability and computational requirements. Our presentation is completed with a discussion of recent trends in deconvolution, inspired by the results of the Grand Challenge on deconvolution microscopy that was recently organized.

J. Icha, Michael Weber, J. Waters et al.

D. Gambarotto, Fabian Zwettler, M. Le Guennec et al.

Determining the structure and composition of macromolecular assemblies is a major challenge in biology. Here we describe ultrastructure expansion microscopy (U-ExM), an extension of expansion microscopy that allows the visualization of preserved ultrastructures by optical microscopy. This method allows for near-native expansion of diverse structures in vitro and in cells; when combined with super-resolution microscopy, it unveiled details of ultrastructural organization, such as centriolar chirality, that could otherwise be observed only by electron microscopy. U-ExM enables near-native expansion microscopy of samples in vitro and in cells. The combination of U-ExM with confocal microscopy and HyVolution revealed details of centriole chirality that were previously accessible only by electron microscopy.

R. Prevedel, A. Diz-Muñoz, G. Ruocco et al.

Martin Schorb, I. Haberbosch, W. Hagen et al.

The demand for high-throughput data collection in electron microscopy is increasing for applications in structural and cellular biology. Here we present a combination of software tools that enable automated acquisition guided by image analysis for a variety of transmission electron microscopy acquisition schemes. SerialEM controls microscopes and detectors and can trigger automated tasks at multiple positions with high flexibility. Py-EM interfaces with SerialEM to enact specimen-specific image-analysis pipelines that enable feedback microscopy. As example applications, we demonstrate dose reduction in cryo-electron microscopy experiments, fully automated acquisition of every cell in a plastic section and automated targeting on serial sections for 3D volume imaging across multiple grids. Py-EM and SerialEM enable automated microscope control for high-throughput data acquisition in diverse transmission electron microscopy imaging experiments.

J. Jonkman, C. Brown, G. Wright et al.

Confocal Microscopy

it should be either heated (+37 °C) or at room temperature for the whole day. The first reservation created for a given day determines the temperature of the system.

A. Gruverman, M. Alexe, D. Meier

Since its inception more than 25 years ago, Piezoresponse Force Microscopy (PFM) has become one of the mainstream techniques in the field of nanoferroic materials. This review describes the evolution of PFM from an imaging technique to a set of advanced methods, which have played a critical role in launching new areas of ferroic research, such as multiferroic devices and domain wall nanoelectronics. The paper reviews the impact of advanced PFM modes concerning the discovery and scientific understanding of novel nanoferroic phenomena and discusses challenges associated with the correct interpretation of PFM data. In conclusion, it offers an outlook for future trends and developments in PFM.

Amicia D Elliott

In light microscopy, illuminating light is passed through the sample as uniformly as possible over the field of view. For thicker samples, where the objective lens does not have sufficient depth of focus, light from sample planes above and below the focal plane will also be detected. The out‐of‐focus light will add blur to the image, reducing the resolution. In fluorescence microscopy, any dye molecules in the field of view will be stimulated, including those in out‐of‐focus planes. Confocal microscopy provides a means of rejecting the out‐of‐focus light from the detector such that it does not contribute blur to the images being collected. This technique allows for high‐resolution imaging in thick tissues.

Seungwan Jeon, Jongbeom Kim, Donghyun Lee et al.

Photoacoustic imaging (PAI) has many interesting advantages, such as deep imaging depth, high image resolution, and high contrast to intrinsic and extrinsic chromophores, enabling morphological, functional, and molecular imaging of living subjects. Photoacoustic microscopy (PAM) is one form of the PAI inheriting its characteristics and is useful in both preclinical and clinical research. Over the years, PAM systems have been evolved in several forms and each form has its relative advantages and disadvantages. Thus, to maximize the benefits of PAM for a specific application, it is important to configure the PAM system optimally by targeting a specific application. In this review, we provide practical methods for implementing a PAM system to improve the resolution, signal-to-noise ratio (SNR), and imaging speed. In addition, we review the preclinical and the clinical applications of PAM and discuss the current challenges and the scope for future developments.

Ke Bian, Christoph Gerber, A. Heinrich et al.

T. Cocker, V. Jelic, R. Hillenbrand et al.

Sudheesh Parathakkatt, Vaisakh Kizhuveetil, Gokul G. K. et al.

Worm-like micelles (WLMs) are dynamic, self-assembling supramolecular structures that exhibit complex viscoelastic behaviour due to their ability to undergo reversible scission, fusion, branching, and sequence rearrangement. This review provides a comprehensive analysis of recent theoretical advances in modelling WLM rheology, from classical reptation–scission theories to modern stochastic simulations and multi-scale population-balance frameworks. A central challenge addressed is the rheological indistinguishability of competing models under linear conditions, which renders inverse modelling ill-posed and necessitates the integration of experimental data, such as cryogenic transmission electron microscopy (cryo-TEM), small-angle neutron scattering (SANS), and flow birefringence, to constrain theoretical predictions. The article further explores the limitations of conventional models in capturing nonlinear responses, including shear banding and extensional strain hardening, and emphasizes the need for spatially resolved, structurally informed constitutive equations. Emerging tools, including neural networks and hybrid modular frameworks, are identified as promising solutions for bridging microscopic rearrangement dynamics with macroscopic flow behaviour. Ultimately, the development of predictive, physically grounded WLM models will be essential for advancing applications in formulation science, smart materials, and industrial processing.

Ernst H. K. Stelzer, Frederic Strobl, Bo-Jui Chang et al.