Hasil untuk "Microscopy"

S. Zheng, Eugene Palovcak, J. Armache et al.

I. Horcas, R. Fernandez, J. Gómez‐Rodríguez et al.

Guang Tang, Liwei Peng, P. Baldwin et al.

A. Spurr

E. Reynolds

Aqueous solutions of lead salts (1, 2) and saturated solutions of lead hydroxide (1) have been used as stains to enhance the electron-scattering properties of components of biological materials examined in the electron microscope. Saturated solutions of lead hydroxide (1), while staining more intensely than either lead acetate or monobasic lead acetate (l , 2), form insoluble lead carbonate upon exposure to air. The avoidance of such precipitates which contaminate surfaces of sections during staining has been the stimulus for the development of elaborate procedures for exclusion of air or carbon dioxide (3, 4). Several modifications of Watson's lead hydroxide stain (1) have recently appeared (5-7). All utilize relatively high pH (approximately 12) and one contains small amounts of tartrate (6), a relatively weak complexing agent (8), in addition to lead. These modified lead stains are less liable to contaminate the surface of the section with precipitated stain products. The stain reported here differs from previous alkaline lead stains in that the chelating agent, citrate, is in sufficient excess to sequester all lead present. Lead citrate, soluble in high concentrations in basic solutions, is a chelate compound with an apparent association constant (log Ka) between ligand and lead ion of 6.5 (9). Tissue binding sites, presumably organophosphates, and other anionic species present in biological components following fixation, dehydration, and plastic embedding apparently have a greater affinity for this cation than lead citrate inasmuch as cellular and extracellular structures in the section sequester lead from the staining solution. Alkaline lead citrate solutions are less likely to contaminate sections, as no precipitates form when droplets of fresh staining solution are exposed to air for periods of up to 30 minutes. The resultant staining of the sections is of high intensity in sections of Aralditeor Epon-embedded material. Cytoplasmic membranes, ribosomes, glycogen, and nuclear material are stained (Figs. 1 to 3). STAIN SOLUTION: Lead citrate is prepared by

Michael J Rust, M. Bates, X. Zhuang

M. Karnovsky, M. Karnovsky, M. Karnovsky et al.

S. Bolte, F. Cordelières

W. Denk, J. Strickler, W. Webb

J. Hobbie, R. Daley, S. Jasper

I. W. Mclean, P. Nakane

M. Gustafsson

S. Hell, Jan Wichmann

G. Zinser

S. Hess, T. P. K. Girirajan, Michael D. Mason

F. Helmchen, W. Denk

R. E. Lee

W. Zipfel, Rebecca M. Williams, W. Webb

Susanne C. M. Reinhardt, Luciano A. Masullo, I. Baudrexel et al.

The authors introduce a single-molecule DNA-barcoding method, resolution enhancement by sequential imaging, that improves the resolution of fluorescence microscopy down to the Ångström scale using off-the-shelf fluorescence microscopy hardware and reagents. Fluorescence microscopy, with its molecular specificity, is one of the major characterization methods used in the life sciences to understand complex biological systems. Super-resolution approaches^ 1 – 6 can achieve resolution in cells in the range of 15 to 20 nm, but interactions between individual biomolecules occur at length scales below 10 nm and characterization of intramolecular structure requires Ångström resolution. State-of-the-art super-resolution implementations^ 7 – 14 have demonstrated spatial resolutions down to 5 nm and localization precisions of 1 nm under certain in vitro conditions. However, such resolutions do not directly translate to experiments in cells, and Ångström resolution has not been demonstrated to date. Here we introdue a DNA-barcoding method, resolution enhancement by sequential imaging (RESI), that improves the resolution of fluorescence microscopy down to the Ångström scale using off-the-shelf fluorescence microscopy hardware and reagents. By sequentially imaging sparse target subsets at moderate spatial resolutions of >15 nm, we demonstrate that single-protein resolution can be achieved for biomolecules in whole intact cells. Furthermore, we experimentally resolve the DNA backbone distance of single bases in DNA origami with Ångström resolution. We use our method in a proof-of-principle demonstration to map the molecular arrangement of the immunotherapy target CD20 in situ in untreated and drug-treated cells, which opens possibilities for assessing the molecular mechanisms of targeted immunotherapy. These observations demonstrate that, by enabling intramolecular imaging under ambient conditions in whole intact cells, RESI closes the gap between super-resolution microscopy and structural biology studies and thus delivers information key to understanding complex biological systems.

Anwai Archit, Sushmita Nair, Nabeel Khalid et al.

We present Segment Anything for Microscopy, a tool for interactive and automatic segmentation and tracking of objects in multi-dimensional microscopy data. Our method is based on Segment Anything, a vision foundation model for image segmentation. We extend it by training specialized models for microscopy data that significantly improve segmentation quality for a wide range of imaging conditions. We also implement annotation tools for interactive (volumetric) segmentation and tracking, that speed up data annotation significantly compared to established tools. Our work constitutes the first application of vision foundation models to microscopy, laying the groundwork for solving image analysis problems in these domains with a small set of powerful deep learning architectures.