Wide-field optically sectioning fluorescence microscopy with laser illumination

We describe an extremely simple method by which optically sectioned fluorescence images may be obtained with conventional microscopes using laser illumination. A one-dimensional grid pattern is introduced into the illumination system, together with a rotating ground glass diffuser. This causes an image of the grid pattern to be projected into the specimen. Images taken at three spatial positions of the grid are processed in a simple manner to provide optically sectioned images of fluorescent specimens.     

Quantitative electron spectroscopic diffraction analyses of the crystal formation in dentine

Newly formed apatitic crystallites of different hard tissues consist, according to our investigations, of chains composed of nanometre-sized particles (islands, dots) arising at nucleating sites of the collagenous and noncollagenous matrix macromolecules. In dentine these islands coalesce rapidly in longitudinal direction to form needle-like crystallites which further coalesce to ribbon-like crystallites. We have concluded that the centre-to-centre distances between these islands represent the distances between the nucleating sites of the matrix macromolecules. We have applied energy-filtering transmission electron microscopy in the selected area electron diffraction mode at different stages of crystal formation in dentine and have obtained quantitative information of the degree of crystal disorder on the basis of the paracrystal theory. The fluctuation of the lattice plane distances in c -axis direction decreases, proceeding from the region near the dentine/predentine border to the dentine/enamel border.     

Uncertainty contribution of tip-sample angle to AFM lateral measurements

AFM measurements are very important for quality control in the photovoltaic, microfluidic, electronic or micro-optic industries. This work proposes an algorithm to complete the uncertainty evaluation of AFM systems along the XY-axis under conditions where tolerance of curved surfaces must be controlled. This algorithm is also tested for tilt angles between tip and sample from 0° to 9° using an experimental arrangement which consists of an AFM instrumented with an inclinometer and four step height standards.Results show good agreement between the theoretical model and experimental results for samples with larger steps TGZ03 (465 nm) and TGZ11 (1416 nm), but with poor results for the smaller samples TGZ01 (17.6 nm) and TGZ02 (73.1 nm). An angle of 9° shows an error of about 3% in the horizontal determination of the step dimension, but it could increase to 47% for a tilt angle of 30° according to the theoretical model.The angle error between tip and sample is included in the uncertainty budget using a uniform distribution. An evaluation is performed in a theoretical rolling machine for imprint lithography where a step must be measured with nominal dimensions of 3 μm—X-axis and 1 μm—Z-axis. An assumed tip-sample angle is assumed that changes from 0° to 22.5° (curved form) and produces an uncertainty contribution to the X measurement of 55.7 nm. This uncertainty is important and must be considered to guarantee tolerances in quality control of curved form products.     

Growth of carbon nanofilaments on coal foams

Nanofilamentous carbon was grown on a carbon foam by catalytic chemical vapour deposition (CVD) using the decomposition of ethylene/hydrogen mixtures over Ni. The carbon foam was obtained from a coal by a two-stage thermal process, with the first stage taking place at a temperature within the plastic region of the precursor coal. The extent of porosity and the pore size of the foam were mainly influenced by the pressure reached in the reactor during the first stage. In the CVD process, 700 °C was the optimum temperature for obtaining good yields of nanofilaments. A low ethylene/hydrogen ratio (1/4) in the reactive gas gave rise to almost only short and thin carbon nanostructures. A higher proportion of C₂H₄ (4/1, C₂H₄/H₂) gave better yields of nanofilaments, with good proportions of higher-length and higher-diameter (up to around 0.5 μm) structures. Among the carbon forms produced, transmission electron microscopy revealed the predominance of fishbone-type nanofibres, with some bamboo-like nanotubes being also observed.     

Apoptotic DNA fragmentation can be revealed in situ: An ultrastructural approach

A common pattern of apoptotic death is DNA cleavage, initially producing large fragments (50 kbp), followed by the production of nucleosomic/oligonucleosomic fragments. Nevertheless, apoptosis without DNA fragmentation, at least of the nucleosomic type, has been reported. To investigate the spatial relationship between DNA cleavage and chromatin condensation, we applied the TUNEL technique to the ultrastructural analysis of apoptotic cells. A modified method, utilizing a gold‐conjugated antidigoxigenin antibody, was carried out on U937 versus Molt‐4 cells, both exposed to UVB radiation or staurosporine treatment. Gold particle density in the different domains of apoptotic cells was evaluated by a four‐way ANOVA test. Gold labelling was more strongly localised in condensed chromatin than in the diffuse chromatin. U937 cells, which evidenced in vitro oligonucleosomic fragmentation after both UVB and staurosporine treatments, revealed a significantly higher gold particle density, when compared with Molt‐4, which did not show, on the other hand, oligonucleosomic cleavage even in the presence of ≤50 kbp cleavage. Thus, a correlation between DNA fragment sizes and gold particle density appears. TUNEL applied to electron microscopy is an effective approach to study the relationship between apoptotic chromatin condensation and DNA cleavage. Both these events indeed appear in the apoptotic nucleus, but their reciprocal correlation is still greatly unknown. Microsc. Res. Tech. 2009. © 2009 Wiley‐Liss, Inc.     

Structural Perspectives on the Mechanism of Soluble Guanylate Cyclase Activation

The enzyme soluble guanylate cyclase (sGC) is the prototypical nitric oxide (NO) receptor in humans and other higher eukaryotes and is responsible for transducing the initial NO signal to the secondary messenger cyclic guanosine monophosphate (cGMP). Generation of cGMP in turn leads to diverse physiological effects in the cardiopulmonary, vascular, and neurological systems. Given these important downstream effects, sGC has been biochemically characterized in great detail in the four decades since its discovery. Structures of full-length sGC, however, have proven elusive until very recently. In 2019, advances in single particle cryo–electron microscopy (cryo-EM) enabled visualization of full-length sGC for the first time. This review will summarize insights revealed by the structures of sGC in the unactivated and activated states and discuss their implications in the mechanism of sGC activation.     

Identification of cheese mite species inoculated on Mimolette and Milbenkase cheese through cryogenic scanning electron microscopy

Samples of Mimolette (France) and Milbenkase (Germany) cheeses traditionally ripened by mites were analyzed to determine the mite species present on each sample. Scientific literature was reviewed to understand which mite species most commonly infest cheese. Morphological features possessed by mites were then studied to understand what unique characteristics are required to ensure accurate identification. After identification and compilation of a detailed key of stored food mites (subclass Acari, order Astigmata) and their delineating features, the mites were viewed through a cryogenic scanning electron microscope. It was determined that Mimolette cheese is inoculated with Acarus siro L. The features studied to identify this mite species included idiosomal length and shape, setae length and arrangement, leg size, placement of anus and genitals, and solenidia shape. The Milbenkase cheese is inoculated with Tyrolichus casei Oudemans, which was evident after viewing the same features used to identify A. siro and the supracoxal seta shape. With this knowledge, further research can be conducted on the 2 cheese varieties to understand what chemical, physical, and microbial changes occur within the cheeses because of mites. It is important to identify the mite species present on each cheese variety to improve our understanding of their role in creating the distinctive characteristics that set these cheeses apart from others.     

Synthesis and nonlinear optical properties of a novel polyurethane containing cyanovinylthiophene with enhanced thermal stability of dipole alignment for electro‐optic applications

Stabilization of electrically induced dipole alignment is one of the important criteria in the development of nonlinear optical (NLO) polymers for electro‐optic device applications. Polyurethanes for NLO applications have attracted attention because of their high thermal stability due to hydrogen bonding. In the work reported here, we designed and synthesized a new type of NLO polyurethane, in which the pendant NLO chromophores are part of the polymer backbone. This mid‐type NLO polymer is expected to have the merits of both main‐chain and side‐chain NLO polymers, namely stable dipole alignment and good solubility. 1‐[3,4‐Di‐(2‐hydroxyethoxy)phenyl]‐2‐(2‐thienyl)ethene was prepared and condensed with 3,3′‐dimethoxy‐4,4′‐biphenylenediisocyanate to yield a polyurethane. This polyurethane was reacted with tetracyanoethylene to give a novel Y‐type polyurethane (7) containing 1‐(3,4‐dioxyphenyl)‐2‐[5‐(1,2,2‐tricyanovinyl)‐2‐thienyl]ethenes as NLO chromophores, which constitute part of the polymer backbone. Polyurethane 7 is soluble in common organic solvents such as N,N‐dimethylformamide and dimethylsulfoxide. It shows a thermal stability up to 280 °C from thermogravimetric analysis with a glass transition temperature obtained from differential scanning calorimetry of ca 162 °C. The second harmonic generation (SHG) coefficient (d₃₃) of a poled polymer film of he polyurethane at 1560 nm fundamental wavelength is ca 1.11 × 10^?18 C. Polymer 7 exhibits an enhanced thermal stability and no significant SHG decay is observed below 150 °C, which is acceptable for NLO device applications. Copyright © 2009 Society of Chemical Industry     

Aqueous lateral epitaxy overgrowth of ZnO on (0001) GaN at 90 °C: Part I. Increasing the critical thickness

Thick, epitaxial ZnO thin films have been grown on (0001) GaN buffered Al₂O₃ substrates using an aqueous solution at 90 °C. Films with improved structural, optical and electrical characteristics, were grown using a lateral epitaxial overgrowth (LEO) method. Different photoresist masks were used to enable LEO. The masks included linear windows and two different hexagonal arrays of circular windows. Films that exceeded a critical thickness mechanically failed through buckling, consistent with the large compressive stresses expected due to the mismatch of the ZnO lattice with the underlying GaN substrate. It was shown that improved mechanical stability could be achieved using the LEO method. Without LEO, a film thickness no greater than 4 µm could be grown without buckling. The critical thickness could be increased to 10 µm using linear windows, whereas a critical thickness of 50 μm was achieved with one array of circular windows, and 80 µm for a second array. The two different arrays of circular windows differed relative to their orientation on the substrate. It was also shown that the critical thickness increased with increasing distance between the growth windows. Optical transmission, micro-photoluminescence and Hall Effect measurements showed that the LEO method also results in improved optoelectronic properties.