Transmission Electron Microscopy Grids Research Articles

Bienzyme liquid-crystal-based cholesterol biosensor

A cholesterol biosensor was fabricated by coating 4-cyano-4′-pentylbiphenyl (5CB) in a transmission electron microscope (TEM) grid with poly(acrylicacid-b-4-cyno-4′-undecyl acrylate) (PAA-b-LCP), followed by co-immobilization of cholesterol oxidase and horseradish peroxidase on the PAA brushes. This TEM grid sensor was tested for stability and sensitivity of the immobilized enzymes to cholesterol. Cholesterol was detected by a planar-to-homeotropic orientational change of 5CB at a limit of 0.8mM. The high stability, ease of preparation, and specific detection of cholesterol in a complex mixture, could provide a new and convenient method for screening cholesterol levels in a sample using a polarized optical microscope.

Erbium doped aluminum nitride nanoparticles for nano-thermometer applications

We have synthesized nanoparticles (NPs) of aluminum nitride (AlN) doped in situ with erbium (Er) using the inert gas condensation technique. These NPs have optical properties that make them good candidates for nanoscale temperature sensors. The photoluminescence (PL) spectrum of Er3+ in these NPs shows two emission peaks in the green region at around 540 and 560 nm. The ratio of the intensities of these luminescence peaks is related to temperature. Using Boltzmann’s distribution, the temperature of the NP and its surrounding can be calculated. The NPs were directly deposited on (111) p-type silicon wafers, transmission electron microscope grids and glass cover slips. XRD and HRTEM study indicates that most of the NPs have crystalline hexagonal AlN structure. An enhancement of the luminescence from these NPs was observed after heating in-air at 770 K for 3 h. The sample was then heated in air using a scanning optical microscope laser. The corresponding change in PL peak intensities of the NPs was recorded for laser powers ranging from 0.2 to 15.1 mW. Temperature calculated using the Boltzmann’s distribution was in the range of 300–470 K. This temperature range is of interest for semiconductor device heating and for thermal treatment of cancerous cells, for example.

A General Technique to Investigate the Aggregation of Nanoparticles by Transmission Electron Microscopy.

The aggregation state of nanoparticles (NPs) must be precisely known in order to study the structure-property relationship and to evaluate the exploitability of NPs dispersion for a given application. Here we report a general technique for sample preparation to investigate with transmission electron microscopy (TEM) the state of aggregation of NPs dispersed in liquid solution. Following a simple procedure which requires few minutes, the aggregates of NPs are "frozen" in a polymeric matrix simultaneously to their deposition on a TEM grid. Our technique is of general applicability and it avoids the use of cryo-TEM, which is more expensive, more time consuming and less common than ordinary TEM. Compared to the investigation of NPs aggregation in the liquid phase with dynamic light scattering, our approach avoids the problem of shielding by large aggregates and it allows the full exploitation of TEM advantages, primarily the reliable determination of shape and size of each aggregate and the precise evaluation of the number of single NPs forming each cluster. As an example, we demonstrate the use of our technique to study two frequent topics related to aggregation: the plasmon properties of gold NPs aggregates and the stability of iron oxide NPs in physiological environment. The methodology described here will be useful to advance the knowledge about how aggregation influences the physical-chemical properties of NPs.

Structural evolution of soot particles during diesel combustion in a single-cylinder light-duty engine

The structural evolution of soot particles in a single-cylinder, light-duty diesel engine has been investigated by conducting thermophoretic soot sampling and subsequent transmission electron microscope (TEM) imaging. The location of TEM grid with respect to a diesel flame is varied so that soot particles are sampled at three different combustion stages including (1) when the impingement of flame on the wall occurs, (2) after the flame impingement, and (3) during the late-cycle burn-out. For comparison purposes, engine-out soot particles are also collected at the same operating conditions. The results show that diesel soot particles are aggregates of varying numbers of primary particles. It is found that the flame impingement on the wall makes a significant impact on the soot aggregate structures, evidenced by the decreased mean radius of gyration of the aggregates from 38 to 26nm between the wall-impinging and post-impingement stages. This was due primarily to the fragmentation of large soot aggregates while the mean diameter of primary particles remains the same. From the post-impingement to late-cycle burn-out stages, most of the soot aggregates disappear due to the oxidation leaving only highly agglomerated substructures. As a result, soot aggregates show higher fractal dimension in the late-cycle burn-out stage than that in the previous stages. The intensive soot oxidation also reduces primary particle diameter from 19 to 15nm through surface oxidation. The engine-out soot samples show similar particle size and structure to the late-cycle samples, suggesting that the late-cycle soot particles experienced little oxidation before exiting through the exhaust. This leads to a conclusion that the highly agglomerated substructures of soot aggregates can survive the late-cycle burn-out and become a major contributor to exhaust soot emissions.

Freestanding macroscopic metal-oxide nanotube films derived from carbon nanotube film templates

Aligned carbon nanotube films coated with amorphous carbon were developed into novel templates by atomic layer deposition. Freestanding macroscopic metal-oxide nanotube films were then successfully synthesized by using these templates. The reactive amorphous carbon layer greatly improved the nuclei density, which ensured the high quality of the films and allowed for precise control of the wall thickness of the nanotubes. Using template-synthesized alumina nanotube films, we demonstrate a humidity sensor with a high response speed, a transmission electron microscopy (TEM) grid, and a catalyst support. The cross-stacked assembly, ultrathin thickness, chemical inertness, and high thermal stability of the alumina nanotube films contributed to the excellent performance of these devices. In addition, it is expected that the metal-oxide nanotube films would have significant potential owing to their material richness, macroscopic appearance, flexibility, compatibility with the semiconducting technologies, and the feasibility of mass production.

Controlled Vortex Formation and Facilitated Energy Transfer within Aggregates of Colloidal CdS Nanorods

Vortex formation is observed in transmission electron microscopy (TEM) images of colloidal CdS nanorod (NR) aggregates. Upon adding polar (poor) solvents into the NR dispersion, the NRs passivated with alkylphosphonic acids start flocculating. When NR clusters are deposited on a TEM grid, the tips of the NRs are immobilized on the substrate, and capillary forces give rise to a net inward tilting as the solvent evaporates, eventually leading to the formation of vortices. Solvent polarity turns out to play a critical role in controlling the structure of vortices. Fluorescence lifetime imaging microscopy (FLIM) of the ensembles reveals that the vortex structures have profound influence on the energy transfer rate within the NR assembly.

A simple but precise method for quantitative measurement of the quality of the laser focus in a scanning optical microscope.

SummaryWe report a method for characterizing the focussing laser beam exiting the objective in a laser scanning microscope. This method provides the size of the optical focus, the divergence of the beam, the ellipticity and the astigmatism. We use a microscopic‐scale knife edge in the form of a simple transmission electron microscopy grid attached to a glass microscope slide, and a light‐collecting optical fibre and photodiode underneath the specimen. By scanning the laser spot from a reflective to a transmitting part of the grid, a beam profile in the form of an error function can be obtained and by repeating this with the knife edge at different axial positions relative to the beam waist, the divergence and astigmatism of the postobjective laser beam can be obtained. The measured divergence can be used to quantify how much of the full numerical aperture of the lens is used in practice. We present data of the beam radius, beam divergence, ellipticity and astigmatism obtained with low (0.15, 0.7) and high (1.3) numerical aperture lenses and lasers commonly used in confocal and multiphoton laser scanning microscopy. Our knife‐edge method has several advantages over alternative knife‐edge methods used in microscopy including that the knife edge is easy to prepare, that the beam can be characterized also directly under a cover slip, as necessary to reduce spherical aberrations for objectives designed to be used with a cover slip, and it is suitable for use with commercial laser scanning microscopes where access to the laser beam can be limited.

High-resolution TEM analysis of flatten carbon nanotube packing in nanocomposites

Due to the nanoscale size of carbon nanotubes (CNTs), observing the CNT/CNT and CNT/resin interactions in nanocomposites requires a high resolution transmission electron microscope (TEM) analysis. This research focuses on developing a protocol to provide an effective and reliable approach to obtain ultra-thin (<25nm) cross sections of CNT/polymer nanocomposites for TEM analysis. Improvements were achieved by studying the relationships between chosen microtome cutting parameters, sample characteristics, and TEM image quality. A cutting protocol was established so that ultra-thin cross sections can be achieved by different microtome operators. In addition, a custom made tool was utilized to aid in the sample collection process. The 3D printed tool made it possible to view both the cut sample and TEM grid under the same optical view, ensuring that the cut sample was successfully transported to the TEM grid.Two CNT nanocomposite samples made with different resins were studied for both microtome cutting and TEM analysis and it revealed the alignment direction of the nanotubes and numerous stacks of CNT bundles. In addition, there was visible flattening of CNT packing into dumbbell shapes similar to results obtain in CNT yarn materials. TEM for the 3-layer CNT BP/BMI nanocomposite revealed uniform cutting in the resin areas. However, when the diamond knife reached graphite crystalline regions, the nanotubes either became deformed into a cone-like structure, were cut thicker than the resin, or folded over onto itself. This is most likely due to the high mechanical properties of CNT in response to the stress of cutting.

Role of Fluid-Dynamics in Soot Formation and Microstructure in Acetylene-Air Laminar Diffusion Flames

Residence time and thermo-chemical environment are important factors in the soot-formation process in flames. Studies have revealed that flow-dynamics plays a dominant role in soot formation process. For understanding the effect of flow dynamics on soot formation and physical structure of the soot formed in different combustion environments two types of laminar diffusion flames of Acetylene and air, a normal diffusion flame (NDF) and an inverse diffusion flame (IDF) have been investigated. The fuel and air supply in the reaction zone in two flame types were kept constant but the interchange of relative position of fuel and air altered the burner exit Reynolds and Froude numbers of gases, fuel/air velocity ratio and flame shape. Soot samples were collected using thermophoretic sampling on transmission electron microscope (TEM) grids at different flame heights and were analyzed off-line in a Transmission Electron Microscope. Soot primary particle size, soot aggregate size and soot volume fraction were measured using an image analysis software. In NDF the maximum flame temperature was about 1525 K and 1230 K for IDF. The soot primary particles are distinctly smaller in size in IDF (between 19 – 26 nm) compared to NDF (between 29–34 nm). Both NDF and IDF show chainlike branched structure of soot agglomerate with soot particles of a nearly spherical shape. The average number of soot primary particles per aggregate in NDF was in the range of 24 to 40 and in IDF it varied between 16 to 24. Soot volume fraction was between 0.6 to 1.5 ppm in NDF where as it was less than 0.2 ppm in IDF. The change in sooting characteristics of the two flame types is attributed to changed fuel/air velocity ratio, entrainment of gas molecules and thermophoresis on soot particles.

Polarized Raman spectroscopy with differing angles of laser incidence on single-layer graphene

Chemical vapor deposition (CVD)-grown single-layer graphene samples, transferred onto a transmission electron microscope (TEM) grid and onto a quartz plate, were studied using polarized Raman spectroscopy with differing angles of laser incidence (θ). Two different polarization configurations are used. In an in-plane configuration, the polarization direction of both incident and scattered light is parallel to the graphene plane. In an out-of-plane configuration, the angle between the polarization vector and the graphene plane is the same as the angle of laser incidence (θ). The normalized Raman intensity of the G-band measured in the out-of-plane configuration, with respect to that in the in-plane configuration, was analyzed as a function of θ. The normalized Raman intensity showed approximately cos2θ-dependence up to θ = 70°, which can be explained by the fact that only the electric field component of the incident and the scattered photon in the out-of-plane configuration projected onto the graphene plane can contribute to the Raman scattering process because of the perfect confinement of the electrons to the graphene plane.

Liquid crystal-based glucose biosensor functionalized with mixed PAA and QP4VP brushes

4-Cyano-4′-pentylbiphenyl (5CB) in a transmission electron microscopy (TEM) grid was developed for glucose detection by coating with a monolayer of mixed polymer brushes using poly(acrylicacid-b-4-cynobiphenyl-4′-oxyundecylacrylate) (PAA-b-LCP) and quaternized poly(4-vinylpyridine-b-4-cynobiphenyl-4′-oxyundecylacrylate) (QP4VP-b-LCP) (LCP stands for liquid crystal polymer) at the 5CB/aqueous interface. The resultant 5CB in TEM grid was functionalized with the PAA and QP4VP brushes, which were strongly anchored by the LCP block. The PAA brush rendered the 5CB/aqueous interface pH-responsive and the QP4VP brush immobilized glucose oxidase (GOx) through electrostatic interactions without the aid of coupling agents. The glucose was detected through a homeotropic-to-planar orientational transition of the 5CB observed through a polarized optical microscope (POM) under crossed polarizers. The optimum immobilization with a 0.78µM GOx solution on the dual-brush-coated TEM grid enabled glucose detection at concentrations higher than 0.5mM with response times shorter than 180s. This TEM grid glucose sensor provided a linear response of birefringence of the 5CB to glucose concentrations ranging from 0.5 to 11mM with a Michaelis–Menten constant (Km) of 1.67mM. This new and sensitive glucose biosensor has the advantages of low production cost, simple enzyme immobilization, high enzyme sensitivity and stability, and easy detection with POM, and may be useful for prescreening the glucose level in the human body.

Morphological characteristics of particles emitted from combustion of different fuels in improved and traditional cookstoves

The study was conducted to analyze the morphological aspects of particle emitted from improved and traditional cookstoves when used with different fuel combinations. The particles were sampled using hood method under actual cooking process performed in controlled laboratory conditions. The sample preparation was done using wet technique which was also compared to dry technique for transferring particles on to the transmission electron microscopy (TEM) grid. The density and parameter fractal dimension 2D-DDf and PDf was calculated using box count algorithm developed using Python language. The 3D-DDf was calculated using ensemble approach and other shape descriptors were also measured using image processing software. The distribution of primary particle, particle agglomerates and tar balls were also studied. The 3D-DDf for particles emitted from traditional cookstove with wood as fuel was 2.42 compared to a lower 3D-DDf of 1.82 in PF cookstove. The 3D-DDf along with fractal prefactor (kg) of 0.7 and 1.4 indicated circular and dense particles emitted from TR cookstove with wood as fuel. On the contrary, with Mix-2 and Mix-3, the particles emitted from improved cookstoves had 3D-DDf>2 indicating presence of super soot aggregates. The distribution of primary particle and tar balls indicated count median diameter (CMD) in the range of 15–25nm and 64–104nm respectively for all cookstove–fuel combinations. The distribution of equivalent area diameter of aggregates showed maximum particles of 300–400nm range as evident in other physical characterization studies conducted using instruments that work light scattering principle and electrical mobility of particles.

Stabilized, Non-Fouling Transmission Electron Microscopy Grid Coatings for the Selective Capture of His-Tag T7 Virus and His-Tag Gro EL from Cell Lysates

Single-particle reconstruction has grown significantly with the improvements in various data collection and computational strategies including CTF fitting, the use of vitrified samples and the utilization of ultra-sensitive direct electron detectors. Although these improvements have contributed significantly to the recent evolution of 3D reconstruction analysis, the way samples are prepared for electron microscopy has remained largely unchanged. We report the development of TEM grids that are modified with a stabilized, non-fouling coating bearing nitrilotriacetic acid substituents at the end of a poly(ethylene glycol)-grafted surface that promotes specific capture of protein targets for single particle reconstruction analysis. The utilization of these grids for specific adsorption of the targeted protein onto the grid surface results in well-controlled surface concentration enhancements and a days-to-minutes reduction in time required for the preparation of a purified sample for cryoEM analysis from an E. coli expression system. The selective and reversible capture of his-tag T7 bacteriophage and GroEL from crude lysates, as well as purified nanodisc-solubilized his-malFGK2, on these NTA-modified grids with an exceptionally low level of adsorption by non-target proteins has been observed. Our data illustrates the utility of these grids for selective capture from complex mixtures, detergent-solubilized membrane protein isolates, and expression systems yielding low copy numbers of the desired target in a manner that is well-suited for single particle reconstruction analysis.

Transmission Electron Microscopy Specimen Preparation for Layer-area Graphene by a Direct Transfer Method

Graphene is a monolayer of carbon atoms arranged in a honeycomb lattice and is a basic building block for graphitic materials of all other dimensionalities. It can be wrapped up into zero-dimensional fullerenes, rolled into one-dimensional nanotubes or stacked into three-dimensional graphite. Graphene has unusual electrical, mechanical, and thermal properties, so it presents new opportunities in fundamental research and practical applications (Zhang et al., 2005; Geim & Novoselov, 2007; Geim, 2009). Recently, large-area synthesis methods for graphene have been advanced epitaxial growth on SiC (Brar et al., 2007; Qi et al., 2010) and chemical vapor deposition on metal substrates such as Ni (Dedkov et al., 2008) and Cu (Li et al., 2009). Particularly, Cu is a considerably attractive substrate because it can be produce layer-area graphene. Transmission electron microscopy (TEM) is a direct and re latively fast imaging tool ideally suited for suspended atomically thin membranes, so it has been successfully applied to study adsorbates on graphene and the atomic structure of graphene (Huang et al., 2011). To observe graphene by TEM, the graphene must transfer from metallic Cu substrates to a TEM grid. Standard transfer of layer graphene has been carried out using a polymer coating such as polymethyl methacrylate (PMMA) or polydimethylsiloxane as a temporary support during etching of the metal substrate to prevent tearing of the graphene (Reina et al., 2009). However, the standard transfer method using these polymers can contaminate and mechanically damage the graphene because it includes several wet chemical steps. Thus, a simple and gentle method is necessary for good TEM images of graphene. Direct transfer method doesn’t include polymer coating process. So it is expected simple and effective method to make graphene specimen by avoiding wet chemical steps (Regan, 2010).

Characterization of exposure to carbon nanotubes in an industrial setting.

While production and use of carbon nanotubes (CNTs) is increasing, workers exposure to CNTs is expected to increase as well, with inhalation being potentially the main pathway for uptake. However, there have been few studies reporting results about workers' personal exposure to CNTs. In this study, worker exposure to single-walled CNTs (SWCNTs) during the production of conductive films in a modern up-scaling factory was assessed. Particulate matter concentrations (2.5-10 μm) and concentrations of CO and CO2 were monitored by using real-time instruments. Workers' exposure levels to SWCNTs were qualitatively estimated by analyzing particle samples by transmission electron microscopy (TEM). TEM samples identified high aspect ratio (length/width > 500) SWCNTs in workplace air. SWCNT concentrations estimated from micrographs varied during normal operation, reactor use without local exhaust ventilation (LEV), and cleaning between 1.7×10(-3), 5.6 and 6.0×10(-3) SWCNT cm(-3), respectively. However, during cleaning it was unclear whether the SWCNTs originated from the cleaning itself or from other reactor openings. We were unable to quantify the SWCNT emissions with online particle instrumentation due to the SWCNT low concentrations compared to background particle concentrations, which were on average 2.6±1.1×10(3)cm(-3). However, CO concentrations were verified as a good indicator of fugitive emissions of SWCNTs. During normal operation, exposure levels were well below proposed limit values (1.0×10(-2) fibers cm(-3) and 1 µg m(-3)) when LEV was used. Based on the results in this study, the analysis of TEM grids seems to be the only direct method to detect SWCNTs in workplace air.

Discotic Liquid Crystal‐Functionalized Gold Nanorods: 2‐ and 3D Self‐Assembly and Macroscopic Alignment as well as Increased Charge Carrier Mobility in Hexagonal Columnar Liquid Crystal Hosts Affected by Molecular Packing and π–π Interactions

Gold nanorods functionalized with triphenylene‐based discotic liquid crystal (LC) motifs show striking self‐assembly behavior both on transmission electron microscopy (TEM) grids as well as in the bulk enforced by the π–π‐stacking of triphenylene groups of adjacent nanorods. TEM images confirm that these discotic LC nanorods form ribbons of parallel‐stacked nanorods several hundred nanometer long. The pursued silane conjugation approach to decorate the nanorods allows for the preparation of dispersions of the nanorods in the hexagonal columnar phases of parent discotic LCs, where the nanorods can be macroscopically aligned with almost 80% efficiency by a simple shearing protocol. Doping the parent host materials with about 1% by weight of the discotic LC‐capped nanorods also reduces the lattice parameter and the intracolumnar packing, which gives rise to enhanced charge carrier mobility in these hosts as determined by time‐of‐flight measurements.

Measurements of the characteristics of particle emissions from a port fuel injection engine

The characteristics of particle emissions from gasoline port fuel injection engines have not been fully investigated and established in comparison with those from direct-injection spark ignition engines. This paper aims to evaluate a new measurement system that could potentially be adapted later to allow a much wider examination of particulate production from port fuel injection engines relative to gasoline direct-injection engines. This work presents the morphology of the particles emitted from port fuel injection engines analysed using scanning transmission electron microscopy. The current designs of dilution tunnel used in particle morphology measurements were reviewed, and subsequently a new dilution tunnel was designed, built, tested and optimised for nanoparticle measurements from port fuel injection engines. A new methodology for capturing nanoparticles from port fuel injection engines for morphological analysis by transmission electron microscopy was investigated by integrating a nanoparticle impactor with a dilution tunnel. The performance of the dilution tunnel demonstrated that it integrated very well with the impactor and achieved the targets in terms of the flow rate and the temperature. The widest variety of particle morphologies, including liquid particles, was observed on the transmission electron microscopy grid from impactor plate A. In addition to soot agglomerates, crystalline particles and salt particles were also seen. In contrast with earlier studies, particle fragmentation was also observed.

Novel lift-off technique for Transmission Electron Microscopy imaging of block copolymer films.

We have developed a simple technique to allow for the lift-off and subsequent transfer of poly(styrene-block-ethylene glycol) films to Transmission Electron Microscopy (TEM) grids. The block copolymer is spin coated onto carbon coated mica and annealed. After the thin film is produced it can easily be floated onto water and picked up by a TEM grid. This method offers better control over film processing than dip coating the TEM grid and is also a significant improvement over methods using etchants such as hydrofluoric acid.

Extreme ultraviolet spectrometer based on a transmission electron microscopy grid

We performed extreme ultraviolet spectroscopy using an 80 lines/mm transmission electron microscope mesh as the dispersive element. We present the usefulness of this instrument for dispersing a high harmonic spectrum from the 13th to the 29th harmonic of a Ti:sapph laser, corresponding to a wavelength range from 60 to 27 nm. The resolution of the instrument is limited by the image size of the high harmonic generation region on the detector. The resolution in first order diffraction is under 2 nm over the entire spectral range with a resolving power around 30.

Phase identification and structure determination from multiphase crystalline powder samples by rotation electron diffraction

Phase identification and structure characterization are important in synthetic and materials science. It is difficult to characterize the individual phases from multiphase crystalline powder samples, especially if some of the phases are unknown. This problem can be solved by combining rotation electron diffraction (RED) and powder X-ray diffraction (PXRD). Four phases were identified on the same transmission electron microscopy grid from a multiphase sample in the Ni–Se–O–Cl system, and their structures were solved from the RED data. Phase 1 (NiSeO3) was found in the Inorganic Crystal Structure Database using the information from RED. Phase 2 (Ni3Se4O10Cl2) is an unknown compound, but it is isostructural to Co3Se4O10Cl2, which was recently solved by single-crystal X-ray diffraction. Phase 3 (Ni5Se6O16Cl4H2) and Phase 4 (Ni5Se4O12Cl2) are new compounds. The fact that there are at least four different compounds in the as-synthesized material explains why the phase identification and structure determination could not be done by PXRD alone. The RED method makes phase identification from such multiphase powder samples much easier than would be the case using powder X-ray diffraction. The RED method also makes structure determination of submicrometre-sized crystals from multiphase samples possible.