31 research outputs found
Three-Dimensional Analysis and Computer Modeling of the Capillary Endothelial Vesicular System with Electron Tomography
No full textObjective: We examined the three-dimensional organization of the endothelial vesicular system with TEM tomography of semi-thick sections. Materials and methods: Mouse abdominal muscle capillaries were perfused with terbium to label vesicular compartments open to the luminal surface. The tissue was prepared for TEM and semi-thick (250nm) sections were cut. Dual axis tilt series, collected from +60° to -60° at 1° increments, were acquired in regions of labeled abluminal caveolae. These tomograms were reconstructed and analyzed to reveal three-dimensional vesicular associations not evident in thin sections. Results: Reconstructed tomograms revealed free vesicles, both labeled and unlabeled, in the endothelial cytoplasm as well as transendothelial channels that spanned the luminal and abluminal membranes. A large membranous compartment connecting the luminal and abluminal surfaces was also present. Computer modeling of tomographic data and video animations provided three-dimensional perspectives to these structures. Conclusions: Uncertainties associated with other three-dimensional methods to study the capillary wall are remedied by tomographic analysis of semi-thick sections. Transendothelial channels of fused vesicles and free cytoplasmic vesicles give credence to their role as large pores in the transport of solutes across the walls of continuous capillaries
Identification of neuromuscular junctions by correlative confocal and transmission electron microscopy
No full textHeterogeneous Extractive Batch Distillation of Chloroform - Methanol – Water : Feasibility and Experiments
Get PDFA novel heterogeneous extractive distillation process is considered for separating the azeotropic mixture chloroform – methanol in a batch rectifying column, including for the first time an experimental validation of the process. Heterogeneous heavy entrainer water is selected inducing an unstable ternary heteroazeotrope and a saddle binary heteroazeotrope with chloroform (ternary diagram class 2.1-2b). Unlike to well-known heterogeneous azeotropic distillation process and thanks to continuous water feeding at the column top, the saddle binary heteroazeotrope chloroform – water is obtained at the column top, condensed and further split into the liquid – liquid decanter where the chloroform-rich phase is drawn as distillate. First, feasibility analysis is carried out by using a simplified differential model in the extractive section for determining the proper range of the entrainer flowrate and the reflux ratio. The operating conditions and reflux policy are validated by rigorous simulation with ProSim Batch Column® where technical features of a bench scale distillation column have been described. Six reproducible experiments are run in the bench scale column matching the simulated operating conditions with two sequentially increasing reflux ratio values. Simulation and experiments agree well. With an average molar purity higher than 99%, more than 85% of recovery yield was obtained for chloroform and methanol
Localization of Fluorescently Tagged Protein to Plasmodesmata by Correlative Light and Electron Microscopy
No full textAuthor response: Stromule extension along microtubules coordinated with actin-mediated anchoring guides perinuclear chloroplast movement during innate immunity
No full textThe effect of fixation on the morphology of the late premolt and early postmolt cuticle of the blue crab, Callinectes sapidus
Get PDFThe dorsal carapace from late premolt (D3) and early postmolt (1 hr) blue crabs (Callinectes sapidus) was used to study the effects of fixation on the morphology of the epicuticle, exocuticle, and hypodermis. Tissues were freshly dissected, quick-frozen in liquid nitrogen, or lyophilized. They were then treated with a standard fixation consisting of sequential fixation in glutaraldehyde, osmium tetroxide, tannic acid and uranyl acetate or with a uranyl acetate primary fixative. Lyophilized-unfixed and lyophilized-rehydrated tissues were also examined. Treatments were viewed using transmission and scanning electron microscopy. Standard fixation preserved both cuticle fibers and cellular elements. Results varied due to the impervious nature of cuticle and the formation of a permeability barrier by 1 hr postmolt. Uranyl acetate fixation greatly improved the contrast of fibers and enhanced the visualization of calcification initiation sites at the epicuticle-exocuticle interface and along the interprismatic septa (IPS), but it did not preserve the hypodermis. The IPS varied in staining intensity among samples, but overall fixation was more uniform than standard fixation. Both treatments revealed a diverse collection of fiber types. Anaglyphs showed that horizontal fibers, oriented parallel to the cuticle surface, rotated in successive planes while vertical fibers were oriented perpendicular to the cuticle surface. Of the vertical fiber types observed, some were associated with pore canals, others were not, and certain ones were restricted to specialized regions of the cuticle. Quick-freezing caused voids to occur between horizontal fibers. Voids were accentuated by lyophilization. The cuticle was more distorted by voids when it was quick-frozen or lyophilized and treated with standard fixation than with uranyl acetate fixation. Unfixed lyophilized samples had a unique tubular morphology that exhibited a lamellar periodicity. Rehydration of lyophilized cuticles reconstituted the fibrous matrix, with all fiber types being preserved except the IPS. Fibers in this treatment appeared most similar to uranyl acetate fixations but were more dispersed. Although the tubular morphology of unfixed, lyophilized samples appears to be an artifact of quick-freezing, it speaks to the highly hydrated nature of the cuticle at the time of ecdysis
Multi-modal registration for correlative microscopy using image analogies
Get PDFCorrelative microscopy is a methodology combining the functionality of light microscopy with the high resolution of electron microscopy and other microscopy technologies for the same biological specimen. In this paper, we propose an image registration method for correlative microscopy, which is challenging due to the distinct appearance of biological structures when imaged with different modalities. Our method is based on image analogies and allows to transform images of a given modality into the appearance-space of another modality. Hence, the registration between two different types of microscopy images can be transformed to a mono-modality image registration. We use a sparse representation model to obtain image analogies. The method makes use of corresponding image training patches of two different imaging modalities to learn a dictionary capturing appearance relations. We test our approach on backscattered electron (BSE) scanning electron microscopy (SEM)/confocal and transmission electron microscopy (TEM)/confocal images. We perform rigid, affine, and deformable registration via B-splines and show improvements over direct registration using both mutual information and sum of squared differences similarity measures to account for differences in image appearance
