Transmission electron microscopy of semiconductor quantum dots

We report on plan-view transmission electron microscopy techniques, by which the size, the actual shape and the strain of a coherent quantum dot in semiconductor heterostuctures can be measured very accurately. The bright-field suppressed-diffraction imaging condition where no strong diffracted beam is excited in the sample provides reliable size measurement. Using suppressed-diffraction imaging condition, the intensity contour in a coherent island is related to the height, and thus the detailed shape and the aspect ratio can be extracted. The strain contrast of a coherent island imaged using an exact two-beam dynamical diffraction condition is useful for strain measurement and the corresponding features is related to the shape of an island. The physical origins and accuracy of interpretation of the image contrast are discussed, using the simulations and experimental examples.     

Visualization and characterization of engineered nanoparticles in complex environmental and food matrices using atmospheric scanning electron microscopy

Imaging and characterization of engineered nanoparticles (ENPs) in water, soils, sediment and food matrices is very important for research into the risks of ENPs to consumers and the environment. However, these analyses pose a significant challenge as most existing techniques require some form of sample manipulation prior to imaging and characterization, which can result in changes in the ENPs in a sample and in the introduction of analytical artefacts. This study therefore explored the application of a newly designed instrument, the atmospheric scanning electron microscope (ASEM), which allows the direct characterization of ENPs in liquid matrices and which therefore overcomes some of the limitations associated with existing imaging methods. ASEM was used to characterize the size distribution of a range of ENPs in a selection of environmental and food matrices, including supernatant of natural sediment, test medium used in ecotoxicology studies, bovine serum albumin and tomato soup under atmospheric conditions. The obtained imaging results were compared to results obtained using conventional imaging by transmission electron microscope (TEM) and SEM as well as to size distribution data derived from nanoparticle tracking analysis (NTA). ASEM analysis was found to be a complementary technique to existing methods that is able to visualize ENPs in complex liquid matrices and to provide ENP size information without extensive sample preparation. ASEM images can detect ENPs in liquids down to 30 nm and to a level of 1 mg L^?1 (9×10⁸ particles mL^?1, 50 nm Au ENPs). The results indicate ASEM is a highly complementary method to existing approaches for analyzing ENPs in complex media and that its use will allow those studying to study ENP behavior in situ, something that is currently extremely challenging to do.     

Interfacial ultramorphology evaluation of resin luting cements to dentin: A correlative scanning electron microscopy and transmission electron microscopy analysis

The objective of this study was to analyze the dentin‐resin cements interfacial ultramorphologies using two different methods: scanning (SEM) and transmission electron microscopy (TEM). Four commercial products were evaluated: two conventional cementing system (RelyX ARC/Adper? Scotchbond? Multi‐Purpose Plus, 3M ESPE and Clearfil Esthetic Cement/DC Bond, Kuraray) and two self‐adhesive resin cements (RelyX Unicem, 3M ESPE and Clearfil SA Cement, Kuraray). Prepolymerized resin disks (Sinfony, 3M ESPE) were cemented on oclusal dentin surfaces of 24 third human molars, simulating the indirect restorations. After 24 h, teeth were sectioned into 0.9‐mm thick slabs and processed for microscopy analyses (SEM or TEM/ n = 3). Qualitative characterization of dentin‐resin cement interface was performed. Hybrid layer formation with long and dense resin tags was observed only for RelyX ARC cementing system. Clearfil Esthetic Cement/DC Bond system revealed few and short resin tags formation, whereas no hybridization and resin tags were detected for self‐adhesive resin cements. Some interfacial regions exhibited that the self‐adhesive resin cements were not bonded to dentin, presenting bubbles or voids at the interfaces. In conclusion, TEM and SEM bonding interface analyses showed ultramorphological variations among resin cements, which are directly related to dental bonding strategies used for each resin cement tested. Microsc. Res. Tech. 76:1234–1239, 2013. © 2013 Wiley Periodicals, Inc.     

Towards the imaging of Weibel–Palade body biogenesis by serial block face‐scanning electron microscopy

Electron microscopy is used in biological research to study the ultrastructure at high resolution to obtain information on specific cellular processes. Serial block face‐scanning electron microscopy is a relatively novel electron microscopy imaging technique that allows three‐dimensional characterization of the ultrastructure in both tissues and cells by measuring volumes of thousands of cubic micrometres yet at nanometre‐scale resolution. In the scanning electron microscope, repeatedly an image is acquired followed by the removal of a thin layer resin embedded biological material by either a microtome or a focused ion beam. In this way, each recorded image contains novel structural information which can be used for three‐dimensional analysis. Here, we explore focused ion beam facilitated serial block face‐scanning electron microscopy to study the endothelial cell–specific storage organelles, the Weibel–Palade bodies, during their biogenesis at the Golgi apparatus. Weibel–Palade bodies predominantly contain the coagulation protein Von Willebrand factor which is secreted by the cell upon vascular damage. Using focused ion beam facilitated serial block face‐scanning electron microscopy we show that the technique has the sensitivity to clearly reveal subcellular details like mitochondrial cristae and small vesicles with a diameter of about 50 nm. Also, we reveal numerous associations between Weibel–Palade bodies and Golgi stacks which became conceivable in large‐scale three‐dimensional data. We demonstrate that serial block face‐scanning electron microscopy is a promising tool that offers an alternative for electron tomography to study subcellular organelle interactions in the context of a complete cell.     

Selective detection of luminescence from semiconductor quantum dots by nanosecond time-gated imaging with a colour-masked CCD detector

Quantum dots are of considerable interest as highly detectable labels with broad absorption, narrow spectral emission and good quantum yields. The luminescence emission has a longer decay time than that of the most common fluorophores, leading to facile rejection of much background emission (such as autofluorescence from biological samples) by means of gated detection. Here, it is shown that a new technique, true‐colour nanosecond time‐gated luminescence imaging, can be used for selective detection of quantum dot luminescence and should prove valuable for multiplexed detection on the basis of both spectral emission profile and luminescence decay time.     

Comparative study of dental enamel loss after debonding braces by analytical scanning electron microscopy (SEM)

Clinical procedures when shear forces are applied to brackets suggest adhesion forces between 2.8 and 10.0 MPa as appropriate. In this study dental enamel was evaluated by scanning electron microscopy (SEM) before and after removing the brackets. Thirty bicuspids (previous prophylaxis) with metallic brackets (Roth Inovation 0.022 GAC), Transbond Plus SEP 3M Unitek adhesive and Transbond XT 3M resin were used. The samples were preserved to 37°C during 24 hr and submited to tangential forces with the Instron Universal machine 1.0 mm/min speed load strength resistance debonding. Also the Adhesive Remanent Index (ARI) test was made, evaluating the bracket base and the bicuspid surface. All the bracket SEM images were processed with AutoCAD to determine the enamel detached area. The average value was 6.86 MPa (SD ± 3.2 MPa). ARI value 1= 63.3%, value 2= 20%, value 3= 13.3% and 33% presented value 0. All those samples with dental enamel loss, presented different situations as fractures, ledges, horizontal, and vertical loss in some cases, and some scratch lines. There is no association between the debonding resistance and enamel presence. Less than half of the remanent adhesive on the dental enamel was present in most of the samples when the ARI test was applied. When the resin area increases, the debonding resistance also increases, and when the enamel loss increases, the resin free metallic area of the bracket base decreases in the debonding.     

Photoluminescence properties of multiple stacked planes of GaN/AlN quantum dots studied by near-field optical microscopy

We have studied the photoluminescence properties of GaN quantum dots with submicrometre lateral resolution by means of near-field scanning optical microscopy. The instrument operated at room temperature and was implemented for near-ultra-violet spectroscopy in the illumination-mode configuration. The analysed sample consisted of several stacked planes of GaN/AlN quantum dots grown by molecular beam epitaxy on Si(111) substrate. The photoluminescence maps showed islands in the micrometre range emitting at different wavelengths, confirming the atomic force microscopy studies on the morphology of similar uncapped samples.     

Cryogenic‐temperature electron microscopy direct imaging of carbon nanotubes and graphene solutions in superacids

Cryogenic electron microscopy (cryo‐EM) is a powerful tool for imaging liquid and semiliquid systems. While cryogenic transmission electron microscopy (cryo‐TEM) is a standard technique in many fields, cryogenic scanning electron microscopy (cryo‐SEM) is still not that widely used and is far less developed. The vast majority of systems under investigation by cryo‐EM involve either water or organic components. In this paper, we introduce the use of novel cryo‐TEM and cryo‐SEM specimen preparation and imaging methodologies, suitable for highly acidic and very reactive systems. Both preserve the native nanostructure in the system, while not harming the expensive equipment or the user. We present examples of direct imaging of single‐walled, multiwalled carbon nanotubes and graphene, dissolved in chlorosulfonic acid and oleum. Moreover, we demonstrate the ability of these new cryo‐TEM and cryo‐SEM methodologies to follow phase transitions in carbon nanotube (CNT)/superacid systems, starting from dilute solutions up to the concentrated nematic liquid‐crystalline CNT phases, used as the ‘dope’ for all‐carbon‐fibre spinning. Originally developed for direct imaging of CNTs and graphene dissolution and self‐assembly in superacids, these methodologies can be implemented for a variety of highly acidic systems, paving a way for a new field of nonaqueous cryogenic electron microscopy.     

Transformation reactions of partially gold-coated bioactive Ca-P glass granules investigated by analytical scanning electron microscopy

In the present work, we have clarified the detail of the surface transformation reactions of bioactive calcium-phosphate (Ca-P) glass granules induced by in vivo implantation in rabbit dorsal muscle sites. To this aim we have compared the behaviour, during the same implantation, between the as-prepared and gold-coated only-on-one-side glass granules. The deposited gold layer enabled us to determine very precisely the initial position of the surface of the glass before the transformation took place. In addition, since the gold layer acts as a diffusion barrier, it allowed the study of the direction and the mechanism of crystal growth which occurred at the glass surface. Lapped and polished cross-sections of the samples were examined by backscattered electron (BSE) imaging and quantitative (with standards) x-ray energy dispersive spectroscopy (EDS) in a scanning electron microscope (SEM). The observations showed the presence of an interlayered structure. Quantitative EDS microanalysis performed by profiling the electron beam across the samples indicated the presence of hydrated calcium phosphate in the external layer, an inhomogeneous silica-rich gel-type layer in the middle layer, and an unaffected original Ca-P glass in the centre. From the comparison with those granules gold-coated on one side, we deduced that the hydrated calcium phosphate layer grew towards the interior of the granules at the expense of the starting glass. A simple model, based on the balance of the concentrations of the elements which have diffused in the different layers, is proposed to explain the contribution of the elements constituting the original glass to the formation of the different layers. This result agrees with the experimental data obtained from image analysis and the microstructural behaviour of this type of glass is discussed.     

Quantitative characterization of the mesothelioma-inducing erionite series minerals by transmission electron microscopy and energy dispersive spectroscopy

Air-collected erionite series minerals from Cappadocia region of Turkey were characterized quantitatively by using transmission electron microscopy (TEM) equipped with energy dispersive spectroscopy (EDS). Field emission scanning electron microscopy aided identification of fibrous minerals. Quantitative characterization guidelines for positive identification of erionites proposed by Dogan and Dogan (2008) was applied and the modified balance error formula (E%<10) and Mg-content test <0.80 were performed for each analysis. Erionite species computation showed that the mineral is erionite-K and a mean chemical formula is proposed based upon the TEM-EDS results. Among the 60 analyses, 11 passed E% test (18.3%), 33 passed Mg-content test (55.0%), and only 3 passed both E% and Mg-content tests (5.0%). This shows difficulty of quantitative characterization of the erionite series minerals. However, as erionite is the most carcinogenic mineral known and is classified by IARC as a Group-I (human) carcinogen, it requires special attention from the mineralogical community to help establish its true mineralogical properties. Quantitatively characterized erionite data are very scarce in literature. Correctly identified erionite mineral types will be useful to medical researchers in their search to find a possible cure for the deadly disease of mesothelioma.     

Cryogenic transmission electron microscopy study: preparation of vesicular dispersions by quenching microemulsions

We previously showed that long‐lived nanoemulsions, seeming initially vesicular, might be prepared simply by diluting and cooling (quenching) warm microemulsions with n‐hexadecane with precooled water. In this paper, we confirm that these systems are vesicular dispersions when fresh, and they can be made with similar structures and compositional dependence using alkanes with chain lengths ranging from octane to hexadecane. The nanostructures of fresh nanoemulsions are imaged with cryogenic transmission electron microscopy (cryo‐TEM). We confirm that water‐continuous microemulsions give simple dispersions of vesicles (sometimes unilamellar), typically less than 100 nm in diameter; these systems can avoid separation for over 2 months. Selected samples were also prepared using halogenated alkanes to create additional contrast in the cryo‐TEM, allowing us to confirm that the oil is located in the observed vesicular structures.     

Direct observation by in situ transmission electron microscopy of the behaviour of IF‐MoS2 nanoparticles during sliding tests: influence of the crystal structure

Direct observation of the behaviour of individual inorganic fullerenes (IF)‐MoS₂ nanoparticles in a sliding interface is essential for the understanding of the influence of the intrinsic characteristics of the nanoparticles on their lubrication mechanisms, when they are used as additives in lubricant oil. In this work, in situ transmission electron microscopy sliding tests were performed on two different types of MoS₂ nanoparticles synthesised by two different methods. It is shown that the IF‐MoS₂ nanoparticles having perfect structure with a high crystalline order and without defects are able to roll and to slide under the combined effect of pressure and shear stress, whereas the IF‐MoS₂ nanoparticles containing many defects exfoliate immediately in the same conditions to deliver MoS₂ layers covering the mating surfaces. A link between these results, the lubrication mechanisms of the nanoparticles and their tribological properties at the macro‐scale was established, proving that the lubrication mechanisms of fullerenes depend on their intrinsic characteristics. Copyright © 2013 John Wiley & Sons, Ltd.     

Mechanisms of cell-cell adhesion identified by immunofluorescent labelling with quantum dots: A scanning near-field optical microscopy approach

Scanning near-field optical microscopy (SNOM) has been employed to simultaneously acquire high-resolution fluorescence images along with shear-force atomic force microscopy from cell membranes. Implementing such a technique overcomes the limits of optical diffraction found in standard fluorescence microscopy and also yields vital topographic information. The application of the technique to investigate cell-cell adhesion has revealed the interactions of filopodia and their functional relationship in establishing adherens junctions. This has been achieved via the selective tagging of the cell adhesion protein, E-cadherin, by immunofluorescence labelling. Two labelling routes were explored; Alexa Fluor 488 and semiconductor quantum dots. The quantum dots demonstrated significantly enhanced photostability and high quantum yield making them a versatile alternative to the conventional organic fluorophores often used in such a study. Analysis of individual cells revealed that E-cadherin is predominantly located along the cell periphery but is also found to extend throughout their filopodia. We have demonstrated that with a fully optimised sample preparation methodology, quantum dot labelling in conjunction with SNOM imaging can be successfully applied to interrogate biomolecular localisation within delicate cellular membranes.     

Identification of novel nonedible oil seeds via scanning electron microscopy for biodiesel production

Exploration of substitute energy feed‐stocks is the much‐debated topic in the scientific society due to increasing power crises and related ecological concerns. As a source of sustainable energy, biodiesel turns out to be the best alternative to petro fuels. In this context, nonedible oil‐producing seeds might be a potential source for biodiesel production owing to their environment‐friendly nature and cost‐effectiveness. The current study, consequently, deals with the investigation and identification of micro‐morphological characters between six novel nonedible oil‐bearing seeds employing scanning electron microscopy as possible biodiesel feed‐stocks. Light microscopic examinations show that seed size varies from 0.3 to 1.3 cm in width and 0.5 to 1.5 cm in. Additionally, a large difference in seed color ranges from dark brown, black, and various shades of light brown was also witnessed. The FFA content of the seeds ranges in 0.3–4.1 mg KOH/g, and the seed oil content fall in 30–65% (w/w) range. SEM‐mediated seed ultrastructure investigations displays greater variation in seed size, shape, color, periclinal wall shape, and sculpturing and so on. All the seeds differ from rounded, ovoid, ovate, oblong, flattened, and elliptical shape. Greater variation in seed wall structure has been seen from angular, entire, irregular, straight, elongated, smooth, and polygonal. The periclinal wall arrangements show alteration from flat, depressed, elevated, smooth, pentagonal, bullate, and coarse seed margins. The results obtained from the current study suggest that scanning electron microscopy could be a beneficial tool in vitalizing the hidden micromorphological characters among various nonedible oil producing seeds, which eventually helps in exploration, correct identification, seed classification, and authentication in future.     

Study of the crystallographic architecture of corals at the nanoscale by scanning transmission X-ray microscopy and transmission electron microscopy

We have investigated the nanotexture and crystallographic orientation of aragonite in a coral skeleton using synchrotron-based scanning transmission X-ray microscopy (STXM) and transmission electron microscopy (TEM). Polarization-dependent STXM imaging at 40-nm spatial resolution was used to obtain an orientation map of the c-axis of aragonite on a focused ion beam milled ultrathin section of a Porites coral. This imaging showed that one of the basic units of coral skeletons, referred to as the center of calcification (COC), consists of a cluster of 100-nm aragonite globules crystallographically aligned over several micrometers with a fan-like distribution and with the properties of single crystals at the mesoscale. The remainder of the skeleton consists of aragonite single-crystal fibers in crystallographic continuity with the nanoglobules comprising the COC. Our observation provides information on the nm-scale processes that led to biomineral formation in this sample. Importantly, the present study illustrates how the methodology described here, which combines HRTEM and polarization-dependent synchrotron-based STXM imaging, offers an interesting new approach for investigating biomineralizing systems at the nm-scale.     

Specimen thickness dependence of hydrogen evolution during cryo‐transmission electron microscopy of hydrated soft materials

The evolution of hydrogen from many hydrated cryo‐preserved soft materials under electron irradiation in the transmission electron microscope can be observed at doses of the order of 1000 e nm^?2 and above. Such hydrogen causes artefacts in conventional transmission electron microscope or scanning transmission electron microscopy (STEM) imaging as well as in analyses by electron energy‐loss spectroscopy. Here we show that the evolution of hydrogen depends on specimen thickness. Using wedge‐shaped specimens of frozen‐hydrated Nafion, a perfluorinated ionomer, saturated with the organic solvent DMMP together with both thin and thick sections of frozen‐hydrated porcine skin, we show that there is a thickness below which hydrogen evolution is not detected either by bubble observation in transmission electron microscope image mode or by spectroscopic analysis in STEM electron energy‐loss spectroscopy mode. We suggest that this effect is due to the diffusion of hydrogen, whose diffusivity remains significant even at liquid nitrogen temperature over the length scales and time scales relevant to transmission electron microscopy analysis of thin specimens. In short, we speculate that sufficient hydrogen can diffuse to the specimen surface in thin sections so that concentrations are too low for bubbling or for spectroscopic detection. Significantly, this finding indicates that higher electron doses can be used during the imaging of radiation‐sensitive hydrated soft materials and, consequently, higher spatial resolution can be achieved, if sufficiently thin specimens are used in order to avoid the evolution of hydrogen‐based artefacts.     

New application of air-drying techniques for studying Ephemeroptera and Plecoptera eggs by scanning electron microscopy

Hexamethyldisilizane (HMDS) and tetramethylsilane are organic compounds that are volatile at ambient temperature and which can therefore be used for air-drying biological samples for SEM studies. The techniques using these compounds provide results that are comparable with those obtained by critical point drying, but which involve a very simple process that saves time and money. Both techniques were applied to SEM studies of Ephemeroptera and Plecoptera eggs in order to assess their suitability as alternative methods to critical point drying for these kinds of biological material. The results show no morphological differences between eggs HMDS air-dried and critical point-dried.     

Antennal fine morphology of the threatened beetle Osmoderma eremita (Coleoptera: Scarabaeidae), revealed by scanning electron microscopy

The aim of this study was to characterize the antennal morphology of Osmoderma eremita, a threatened scarab beetle inhabiting tree hollows. O. eremita males produce a sex pheromone, (R)‐(+)‐γ‐decalactone, responsible mainly for the attraction of females but also other males. Gross and fine morphology of microstructures including sensilla, microsculpture and pores were analyzed using Scanning Electron Microscopy. The antenna of O. eremita showed the typical lamellicorn shape of scarab beetles, with a basal scape, a pedicel, a funicle composed of five antennomeres and a club composed of three lamellae. Six different subtypes of sensilla chaetica (Ch.1 ? 6), Böhm sensilla (Bo), one subtype of sensilla basiconica (Ba.1), two subtypes of sensilla coeloconica (Co.1 ? 2), two subtypes of sensilla placodea (Pl.1 ? 2), pores and peculiar folds were described. The two sexes did not show any significant differences in the occurrence and number of the sensilla placodea, known to be responsible for the pheromone reception. Instead, some sexual differences were found on the occurrence and topology of three different microstructures: (1) one subtype of sensillum chaeticum (Ch.2) occurring on the pedicel only in males; (2) a characteristic pore occurring on the funicle only in males; (3) a peculiar fold occurring on different antennomeres of the funicle in the two sexes, on the fourth in males and on the fifth in females. A comparison between sensilla of O. eremita and those of other Scarabaeoidea is provided. Microsc. Res. Tech. 79:178–191, 2016. © 2016 Wiley Periodicals, Inc.     

Study of the atypical formations in the corrosion bulks of an ancient bronze shield,by optical and electron microscopy

This article presents the atypical formations in the structure of the corrosion crust and in the partially mineralized metallic core, which resulted during the underground stay of a bronze shield, dated between the 1st century B.C. and the 1st century A.D. For our study, we choose a representative fragment from the rim of the shield, which was analyzed by optical microscopy and by electron microscopy coupled with energy‐dispersive X‐ray spectroscopy, to study its morphology, its composition, and the location of chemical compounds on the surface and inside the bulk formed during the underground stay, by processes of chemical and physical alteration, assisted by contamination with structural elements from the site. Those processes, by monolithization and mineralization formed a series of structures consisting of congruent elements and phases with a complex composition. Those formations, defined as surface effects generated by exogenous factors and endogenous factors inside the bulk, are frequently found in ancient bronze objects (such as the exterior flat mole formations and the Liesegang effect in the stratigraphic structure of the bulk). Some of those structures have atypical characteristics as regards their structure, composition, and formation mechanism, which may be used in archeometry. Moreover, that includes the object in the category of special cases, in terms of artifact evolution during underground stay and of the atypical formations resulted from the action of pedological and environmental factors. Microsc. Res. Tech. 2012. © 2012 Wiley Periodicals, Inc.     

The use of a hybrid diffraction unit in the measurement of dislocation densities by electron microscopy

The expression used in the electron microscopical measurement of dislocation densities is ?=2N/Lt, where N is the number of intersections between the dislocation lines in a TEM image, L the total length of a set of sampling lines superimposed upon that image (corrected for magnification) and t the thickness of the thin foil containing the dislocation structure. During the estimation of ? it is therefore important to reduce errors in the measurements of all three variables. It is shown how calibration of the microscope magnification, careful consideration of diffraction conditions and use of the hollow cone and rocking beam facilities on the hybrid diffraction unit fitted to the Philips EM 400T can assist in increasing the speed and accuracy of the measurement procedure.