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"Microscopy."
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Correlative 3D superresolution fluorescence and electron microscopy reveal the relationship of mitochondrial nucleoids to membranes
Microscopic images of specific proteins in their cellular context yield important insights into biological processes and cellular architecture. The advent of superresolution optical microscopy techniques provides the possibility to augment EM with nanometer-resolution fluorescence microscopy to access the precise location of proteins in the context of cellular ultrastructure. Unfortunately, efforts to combine superresolution fluorescence and EM have been stymied by the divergent and incompatible sample preparation protocols of the two methods. Here, we describe a protocol that preserves both the delicate photoactivatable fluorescent protein labels essential for superresolution microscopy and the fine ultrastructural context of EM. This preparation enables direct 3D imaging in 500- to 750-nm sections with interferometric photoactivatable localization microscopy followed by scanning EM images generated by focused ion beam ablation. We use this process to \"colorize\" detailed EM images of the mitochondrion with the position of labeled proteins. The approach presented here has provided a new level of definition of the in vivo nature of organization of mitochondrial nucleoids, and we expect this straightforward method to be applicable to many other biological questions that can be answered by direct imaging.
Journal Article
Microscopy : a very short introduction
Using light, electrons, or X-rays, microscopes today form a vital tool not only in biology but in many other disciplines, including materials science and nanotechnology. In this Very Short Introduction Terence Allen describes the scientific principles behind the main forms of microscopy, and the exciting new developments in the field. Beginning with a brief history of microscopy, Allen surveys the diverse and powerful forms of miscroscopes available today, illustrating how microscopy impinges on almost every aspect of our daily lives.
Book
Advanced imaging techniques for assessment of structure, composition and function in biofilm systems
Scientific imaging represents an important and accepted research tool for the analysis and understanding of complex natural systems. Apart from traditional microscopic techniques such as light and electron microscopy, new advanced techniques have been established including laser scanning microscopy (LSM), magnetic resonance imaging (MRI) and scanning transmission X-ray microscopy (STXM). These new techniques allow in situ analysis of the structure, composition, processes and dynamics of microbial communities. The three techniques open up quantitative analytical imaging possibilities that were, until a few years ago, impossible. The microscopic techniques represent powerful tools for examination of mixed environmental microbial communities usually encountered in the form of aggregates and films. As a consequence, LSM, MRI and STXM are being used in order to study complex microbial biofilm systems. This mini review provides a short outline of the more recent applications with the intention to stimulate new research and imaging approaches in microbiology.
Journal Article
High-order superlattices by rolling up van der Waals heterostructures
Two-dimensional (2D) materials
1
,
2
and the associated van der Waals (vdW) heterostructures
3
–
7
have provided great flexibility for integrating distinct atomic layers beyond the traditional limits of lattice-matching requirements, through layer-by-layer mechanical restacking or sequential synthesis. However, the 2D vdW heterostructures explored so far have been usually limited to relatively simple heterostructures with a small number of blocks
8
–
18
. The preparation of high-order vdW superlattices with larger number of alternating units is exponentially more difficult, owing to the limited yield and material damage associated with each sequential restacking or synthesis step
8
–
29
. Here we report a straightforward approach to realizing high-order vdW superlattices by rolling up vdW heterostructures. We show that a capillary-force-driven rolling-up process can be used to delaminate synthetic SnS
2
/WSe
2
vdW heterostructures from the growth substrate and produce SnS
2
/WSe
2
roll-ups with alternating monolayers of WSe
2
and SnS
2
, thus forming high-order SnS
2
/WSe
2
vdW superlattices. The formation of these superlattices modulates the electronic band structure and the dimensionality, resulting in a transition of the transport characteristics from semiconducting to metallic, from 2D to one-dimensional (1D), with an angle-dependent linear magnetoresistance. This strategy can be extended to create diverse 2D/2D vdW superlattices, more complex 2D/2D/2D vdW superlattices, and beyond-2D materials, including three-dimensional (3D) thin-film materials and 1D nanowires, to generate mixed-dimensional vdW superlattices, such as 3D/2D, 3D/2D/2D, 1D/2D and 1D/3D/2D vdW superlattices. This study demonstrates a general approach to producing high-order vdW superlattices with widely variable material compositions, dimensions, chirality and topology, and defines a rich material platform for both fundamental studies and technological applications.
A simple but flexible technique based on a capillary-force-driven rolling-up process produces high-order van der Waals superlattices that are hard to produce with existing fabrication techniques.
Journal Article
Interferometric fluorescent super-resolution microscopy resolves 3D cellular ultrastructure
Understanding molecular-scale architecture of cells requires determination of 3D locations of specific proteins with accuracy matching their nanometer-length scale. Existing electron and light microscopy techniques are limited either in molecular specificity or resolution. Here, we introduce interferometric photoactivated localization microscopy (iPALM), the combination of photoactivated localization microscopy with single-photon, simultaneous multiphase interferometry that provides sub-20-nm 3D protein localization with optimal molecular specificity. We demonstrate measurement of the 25-nm microtubule diameter, resolve the dorsal and ventral plasma membranes, and visualize the arrangement of integrin receptors within endoplasmic reticulum and adhesion complexes, 3D protein organization previously resolved only by electron microscopy. iPALM thus closes the gap between electron tomography and light microscopy, enabling both molecular specification and resolution of cellular nanoarchitecture.
Journal Article
Free-standing homochiral 2D monolayers by exfoliation of molecular crystals
Two-dimensional materials with monolayer thickness and extreme aspect ratios are sought for their high surface areas and unusual physicochemical properties
1
. Liquid exfoliation is a straightforward and scalable means of accessing such materials
2
, but has been restricted to sheets maintained by strong covalent, coordination or ionic interactions
3
–
10
. The exfoliation of molecular crystals, in which repeat units are held together by weak non-covalent bonding, could generate a greatly expanded range of two-dimensional crystalline materials with diverse surfaces and structural features. However, at first sight, these weak forces would seem incapable of supporting such intrinsically fragile morphologies. Against this expectation, we show here that crystals composed of discrete supramolecular coordination complexes can be exfoliated by sonication to give free-standing monolayers approximately 2.3 nanometres thick with aspect ratios up to approximately 2,500:1, sustained purely by apolar intermolecular interactions. These nanosheets are characterized by atomic force microscopy and high-resolution transmission electron microscopy, confirming their crystallinity. The monolayers possess complex chiral surfaces derived partly from individual supramolecular coordination complex components but also from interactions with neighbours. In this respect, they represent a distinct type of material in which molecular components are all equally exposed to their environment, as if in solution, yet with properties arising from cooperation between molecules, because of crystallinity. This unusual nature is reflected in the molecular recognition properties of the materials, which bind carbohydrates with strongly enhanced enantiodiscrimination relative to individual molecules or bulk three-dimensional crystals.
Sonication of layered metallacycle crystals gives free-standing nanosheets held together by weak non-covalent interactions, with chiral surfaces that show improved binding and enantiodiscrimination compared with individual metallacycle molecules.
Journal Article