Visualization of micromorphology of leaf epicuticular waxes of the rubber tree Ficus elastica by electron microscopy

Ultrastructural aspects of leaf epicuticular waxes were investigated in Ficus elastica by scanning and transmission electron microscopy. Glossy leaves of the rubber tree were collected and subjected to different regimes of specimen preparation for surface observations. F. elastica leaves were hypostomatic and stomata were surrounded with a cuticular thickening that formed a rim. The most prominent epicuticular wax structures of F. elastica leaves included granules and platelets. Without fixation and metal coating, epicuticular wax structures could be discerned on the leaf surface by low-vacuum (ca. 7Pa) scanning electron microscopy. In terms of delineation and retention of the structures, the combination of vapor fixation by glutaraldehyde and osmium tetroxide with subsequent gold coating provided the most satisfactory results, as evidenced by high resolution and sharp protrusions of epicuticular waxes. However, erosion of epicuticular wax edges was noted in the immersion fixed leaves, showing less elongated platelets, less distinct wax edges, and granule cracking. These results suggest that the vapor fixation procedure for demonstrating three-dimensional epicuticular wax structures would facilitate characterization of diverse types of waxes. Instances were noted where epicuticular waxes grew over neighboring epidermal ridges and occluded stomata. In cross sections, epicuticular waxes were observed above the cuticle proper and ranged approximately from 100nm to 500nm in thickness. The native leaf epicuticular waxes had many layers of different electron density that were oriented parallel to each other and parallel or perpendicular to the cuticle surface, implying strata of crystal growth. Such retention of native epicuticular wax structures could be achieved through the use of acrylic resin treated with less harsh dehydrants and mild heat polymerization, alleviating wax extraction during specimen preparations.     

Nanostructure of epicuticular plant waxes: Self-assembly of wax tubules

A “surface science approach” is used to investigate the growth process of tubular wax structures on plant leaves: natural epicuticular wax from nasturtium leaves, mainly consisting of (S)-nonacosan-10-ol, has been recrystallized on artificial substrates of different structure and polarity, namely HOPG, glass and silicon oxide, and the growth process and structure have been studied with scanning electron microscopy (SEM) and atomic force microscopy (AFM). As a result the tubular structure as found on the living leaves is reproduced independent of the nature of the substrate and the way of wax deposition. Prerequisite for tubule formation, however, is the admixture of a few percent of alkyl-diols to the nonacosan-10-ol as shown with artificial mixtures of both components.     

Transmission electron microscopy of heteroepitaxial layer structures

The characterization of heterostructural layers by transmission electron microscopy using cleaved wedge specimens proves to be a fast analysis method. Examples are given for GaAlAs/GaAs and strained GaInAs/GaAs layer systems. It is demonstrated that the (200) dark-field contrast of GaInAs/GaAs layers reverses at an In concentration of x ≈ 0.47. Experimental high-resolution electron images of the edges of cleaved 90° wedges compare very well with computer-simulated images. Characteristic image features in GaAs and AlAs are explained by non-linear beam interactions and are shown to be sensitive to electron beam misalignment. These investigations are important to make a more precise assessment of interfaces possible.     

Magnetic stray fields of patterned permalloy structures investigated by photoemission electron microscopy

Using a photoemission electron microscope we determined magnetic stray fields at the edges of permalloy (Ni₈₀Fe₂₀) particles. X-ray magnetic dichroism was used for visualization of magnetic domains. The values of the stray fields were deduced from the deflection of electrons in the image due to the Lorentz force. The stray fields are responsible for the magnetic interaction of adjacent particles with distances much larger than the thickness. The measured magnetic stray field is about 0.023 T for rectangular particles with a thickness of 30 nm and lateral sizes of tens of microns. PACS 68.37.Xy; 75.40.Cx; 75.75.+a     

Lattice imaging of close-packed structures by high resolution electron microscopy: ABO3 perovskite polytypes

Lattice imaging technique of high resolution electron microscopy has been employed to examine 4H, 6H and 9R ABO₃ perovskite polytypes. The lattice images can be correlated with the lattice periodicity and the stacking sequence of AO₃ layers and BO₆ octahedra. The study shows the utility and validity of the lattice imaging technique for the study of relatively close-packed systems. Commonwealth Visiting Professor, University of Oxford (1974–75).     

The hydrophobic coatings of plant surfaces: epicuticular wax crystals and their morphologies, crystallinity and molecular self-assembly

Plant surfaces are the interfaces of the organisms with respect to their environment. In the micro-dimension they show an enormous variety of functional three-dimensional structures. Their materials and structures developed over millions of years by evolutionary processes in which their functionality has been proven and selected by environmental pressures. As a result, nature developed highly functional materials with several amazing properties like superhydrophobicity and superhydrophilicity. These functional structures are built up by a complex biopolymer called cuticle. The cuticle is mainly composed of a three-dimensional network of cutin, and integrated and superimposed lipids called "waxes". Superimposed waxes are also called "epicuticular waxes". Epicuticular waxes often form two- and three-dimensional structures, in dimensions between hundreds of nanometers and some micrometers, which influence the wettability, self-cleaning behaviour and the light reflection at the cuticle interface. This review gives a brief introduction into the functions of the plant epicuticular waxes and summarises the current knowledge about their morphologies, crystal structures, growth by self-assembly and provides an overview about the microscopy and preparation techniques for their analysis.     

Quantitative atomic-scale analysis of interface structures: transmission electron microscopy and local density functional theory

Transmission electron microscopy (TEM) and local density functional theory (LDFT) are combined to analyze the microscopic structure of the rhombohedral twin interface in alpha-Al2O3. LDFT provides interfacial energetics and atomic and electronic structures for three competing models. With high-resolution TEM the atomic structure at the interface is imaged quantitatively along two orthogonal zone axes. Electron energy loss spectroscopy in TEM with nanoscale spatial resolution yields the interfacial electronic structure. Both experiments confirm the theoretically preferred model quantitatively.     

Fracture behaviour of plant epicuticular wax crystals and its role in preventing insect attachment: a theoretical approach

It is known that attachment ability of insects depends among others on the texture of the substrate. On plant surfaces, covered with microscopic epicuticular wax crystals, insect attachment was found to be highly reduced for many insect species. In some plants, this effect may be explained by the contamination of insect adhesive organs (pads) by wax crystals. In the present study, mechanics of the wax crystals fracture during contact formation between insect adhesive pads and plant surface is examined, in order to explain the observed contamination of pads by wax. It is shown that mechanisms of the wax crystal fracture may be rather different, depending on the slenderness ratio of the crystals. Crystals with high values of the ratio may buckle elastically or in an elastic-plastic way. For five plant species under consideration, the critical value of the slenderness ratio for elastic buckling is 26.5, while for elastic-plastic buckling it is 18.7. If the values of the slenderness ratio are lower than the critical values, one has to consider bending of the crystals under the weight of insects. Although this study considered only crystals of a tubular shape, the general approach is valid also for crystals of other shapes.     

Characterization of periodic domain structures in lithium niobate crystals by scanning electron microscopy and X-ray diffraction analysis

Congruent lithium niobate crystals with periodic domain structures formed by the method of thermoelectric postgrowth treatment have been investigated. Periodic domain structures in the samples of polar ZY-, YZ-, and YX-cuts were characterized by scanning electron microscopy, high resolution x-ray diffractometry, and topography. The evaluation and comparison of the secondary electron (SE) signal cycling, electron emission coefficient, and charge value to equalize the differences in the SE signal have allowed us to specify the features of ZY-and YZ-cut structures. A correlation between the crystal lattice distortions near domain boundaries and the type of domain walls has been found. The domain walls separating the areas with the tail-to-tail P _s vectors not only cause stronger crystal lattice distortions near them, but also are charged less quickly under electron irradiation.     

Low energy electron microscopy and photoemission electron microscopy investigation of graphene

Low energy electron microscopy (LEEM) and photoemission electron microscopy (PEEM) are two powerful techniques for the investigation of surfaces, thin films and surface supported nanostructures. In this review, we examine the contributions of these microscopy techniques to our understanding of graphene in recent years. These contributions have been made in studies of graphene on various metal and SiC surfaces and free-standing graphene. We discuss how the real-time imaging capability of LEEM facilitates a deeper understanding of the mechanisms of dynamic processes, such as growth and intercalation. Numerous examples also demonstrate how imaging and the various available complementary measurement capabilities, such as selected area or micro low energy electron diffraction (μLEED) and micro angle resolved photoelectron spectroscopy (μARPES), allow the investigation of local properties in spatially inhomogeneous graphene samples.     

石墨烯的透射电子显微学研究

石墨烯(Graphene)是近几年迅速发展起来的研究热点材料之一,利用透射电子显微镜(TEM)研究Gra-phene的结构特征和原子动态过程,是Graphene研究的重要进展,文章评述了利用透射电子衍射方法对Graphene的层数、堆垛方式、取向和表面形貌等结构特征进行的研究工作,介绍了利用高分辨透射电子显微术在Graphene的表面缺陷、边缘结构及吸附原子等研究领域取得的最新结果.     

石墨烯的透射电子显微学研究

石墨烯（Graphene）是近几年迅速发展起来的研究热点材料之一.利用透射电子显微镜（TEM）研究Graphene的结构特征和原子动态过程,是 Graphene研究的重要进展.文章评述了利用透射电子衍射方法对Graphene的层数、堆垛方式、取向和表面形貌等结构特征进行的研究工作,介绍了利用高分辨透射电子显微术在Graphene的表面缺陷、边缘结构及吸附原子等研究领域取得的最新结果.     

Direct observations of La ordering and domain structures in La0.61Li0.17TiO3 by high resolution electron microscopy

La_0.61Li_0.17TiO₃ microstructures have been studied by high resolution electron microscopy. A local lattice distortion occurs in the vicinity of the domain boundary region due to the twin with an angle of 89°. The average domain size of La_0.61Li_0.17TiO₃ is greater than 20 nm. The domain size and structures of La_0.61Li_0.17TiO₃ differ greatly from those of La-poor compounds, such as La_0.55Li_0.35TiO₃. At a nanoscopic level, microdomains of 20–100 nm in size construct a two-dimensional structure in La-rich compounds, while microdomains of 5–10 nm in size construct a three-dimensional structure in La-poor compounds. In addition, the Li-ion conduction mechanisms for La-rich and La-poor compounds are two- and three-dimensional, respectively.     

Solving modulated structures by X-ray and electron crystallography

X-ray diffraction can be used for accurately determining not only classical, ordinary structures, but also modulated ones. For structures with weak modulations, the modulation induced satellite reflections are often hard to be observed by X-ray diffraction, but they appear clearly in electron diffraction. In these cases, X-ray diffraction will give only average structures whereas electron diffraction will yield information about the modulations. Sr(1.4)Ta(0.6)O(2.9) is a complex modulated compound with weak modulation and small modulated domains. Here we demonstrate the power of combining X-ray and electron crystallography for studying modulated structures on powders. The modulations of Sr(1.4)Ta(0.6)O(2.9) were determined from electron diffraction (SAED) and high resolution electron microscopy (HREM) images. With specially developed image processing techniques, the weak modulations were enhanced, facilitating the interpretation of HREM images in terms of atomic structure.     

Interaction of doxorubicin with the subcellular structures of the sensitive and Bcl-xL-overexpressing MCF-7 cell line: Confocal and low-energy-loss transmission electron microscopy

The present investigation was directed to examine the interaction of the anti-cancer agent doxorubicin (DOX) with the subcellular compartments of the drug sensitive and Bcl-xL-overexpressing (Bcl-xL) MCF-7 cells using confocal and low-energy-loss transmission electron microscopy (LELTEM). Intracellular detection of DOX with LELTEM was carried out without specific antibodies or heavy metal stains but via the electron-induced molecular orbital excitation of the drug. Cells were incubated with 10 μM DOX for 1 min, 1, 24, and 48 h and then examined live by confocal microscope and as very thin sections in an electron microscope equipped with an energy filter having an energy resolution of 1 eV. Ultrastructural localization of DOX, obtained from pairs of images taken at energy losses of 3 ± 1 and 10 ± 1 eV, were analyzed and correlated with the confocal observations. When the sensitive and Bcl-xL cells were examined under the confocal microscope after 1 min, DOX uptake could not be detected in the nuclei nor in the cytoplasm whereas LELTEM observation revealed that at this stage of incubation the drug has already been incorporated by both cell types and that the nuclear membrane, nucleolus, and mitochondria of the Bcl-xL cells were temporally less DOX-responsive as compared to the sensitive cells. As the incubation time increased, nuclear membranes and nucleoli of both cell types appeared equally sensitive to DOX, nonetheless, mitochondria of the Bcl-xL cells remained invulnerable to DOX for 24 h. The results point to LELTEM feasibility to better characterize yet unresolved cellular events caused by DOX and suggest a transitory role for Bcl-xL overexpression in protecting the cellular compartments from DOX invasion.     

Secondary electron microscopy and transmission electron microscopy studies of carbon nanotubes in C-Ni films

Carbon nanotubes films have been studied with SEM and TEM. The studied films were obtained using a two step method: PVD process and CVD process. Strongly defected and curled carbon nanotubes containing Ni nanoparticles formed the film with thickness of about 300–400 nm. Observed carbon nanotubes were of lengths from 100 nm to 300 nm and did not stick to each other.     

Formation and structures of B36N36 and Y@B36N36 clusters studied by high-resolution electron microscopy and mass spectrometry

High-resolution electron microscopy, mass spectrometry and molecular mechanics/orbital calculations of the boron nitride-based clusters showed the formation of B₃₆N₃₆ and Y@B₃₆N₃₆. Image simulations of these clusters confirmed the proposed structure model.     

Characterization of LiBC by phase-contrast scanning transmission electron microscopy

LiBC was used as a model compound for probing the applicability of phase-contrast (PC) imaging in an aberration-corrected scanning transmission electron microscope (STEM) to visualize lithium distributions. In the LiBC structure, boron and carbon are arranged to hetero graphite layers between which lithium is incorporated. The crystal structure is reflected in the PC-STEM images recorded perpendicular to the layers. The experimental images and their defocus dependence match with multi-slice simulations calculated utilizing the reciprocity principle. The observation that a part of the Li positions is not occupied is likely an effect of the intense electron beam triggering Li displacement.