Immunogold FIB‐SEM: Combining Volumetric Ultrastructure Visualization with 3D Biomolecular Analysis to Dissect Cell–Environment Interactions

Volumetric imaging techniques capable of correlating structural and functional information with nanoscale resolution are necessary to broaden the insight into cellular processes within complex biological systems. The recent emergence of focused ion beam scanning electron microscopy (FIB‐SEM) has provided unparalleled insight through the volumetric investigation of ultrastructure; however, it does not provide biomolecular information at equivalent resolution. Here, immunogold FIB‐SEM, which combines antigen labeling with in situ FIB‐SEM imaging, is developed in order to spatially map ultrastructural and biomolecular information simultaneously. This method is applied to investigate two different cell–material systems: the localization of histone epigenetic modifications in neural stem cells cultured on microstructured substrates and the distribution of nuclear pore complexes in myoblasts differentiated on a soft hydrogel surface. Immunogold FIB‐SEM offers the potential for broad applicability to correlate structure and function with nanoscale resolution when addressing questions across cell biology, biomaterials, and regenerative medicine.     

Nonequilibrium microstructures and their evolution in a Fe–Cr–W–Ni–C laser clad coating

The laser-solidified microstructural and compositional characterization and phase evolution during tempering at 963 K were investigated using an analytical transmission electron microscope with energy dispersive X-ray analysis. The cladded alloy, a powder mixture of Fe, Cr, W, Ni, and C with a weight ratio of 10:5:1:1:1, was processed with a 3 kW continuous wave CO₂ laser. The processing parameters were 16 mm/s beam scanning speed, 3 mm beam diameter, 2 kW laser power, and 0.3 g/s feed rate. The coating was metallurgically bonded to the substrate, with a maximum thickness of 730 μm, a microhardness of about 860 Hv and a volumetric dilution ratio of about 6%. Microanalyses revealed that the cladded coating possessed the hypoeutectic microstructure comprising the primary dendritic γ-austenite and interdendritic eutectic consisted of γ-austenite and M₇C₃ carbide. The γ-austenite was a non-equilibrium phase with extended solid solution of alloying elements and a great deal of defect structures, i.e. a high density of dislocations, twins, and stacking faults existed in γ phase. During high temperature aging, in situ carbide transformation occurred of M₇C₃ to M₂₃C₆ and M₆C. The precipitation of M₂₃C₆, MC and M₂C carbides from austenite was also observed.     

Synthesis of sodium iron silicate(NaFe(III)[SiO3]2) nanorods and electrochemical characterization

Sodium iron silicate (NaFe(III)[SiO₃]₂) nanorods have been synthesized using iron nitrate (Fe(NO₃)₃) and sodium silicate (Na₂SiO₃) solutions by means of hydrothermal method. The mixture solution is processed in hydrothermal autoclave first at 180-200 °C for two days and then dried at 70 °C to obtain nanotructured NaFe(III)[SiO₃]₂. It was revealed that NaFe(III)[SiO₃]₂ nanorod with the average diameter of ~ 15 nm and length of several hundreds nm was confirmed by X-ray power diffraction (XRD), scanning electron microscopy (SEM) and transmission electron microscopy (TEM). Cyclic voltammeter (CV) and electrochemical impedance spectra (EIS) investigations show that this kind of NaFe(III)[SiO₃]₂ nanostructures have evident and stable electrochemical redox behavior between potential range of − 0.1-0.55 V in alkaline solution.     

Fabrication of 3D chitosan–hydroxyapatite scaffolds using a robotic dispensing system

A new robotic desktop rapid prototyping (RP) system was designed to fabricate scaffolds for tissue engineering applications. The experimental setup consists of a computer-guided desktop robot and a one-component pneumatic dispenser. The dispensing material (chitosan and chitosan–hydroxyapatite (HA) dissolved in acetic acid) was stored in a 30-ml barrel and forced out through a small Teflon-lined nozzle into a dispensing medium (sodium hydroxide–ethanol in ratio of 7:3). Layer-by-layer, the chitosan was fabricated with a preprogramed lay-down pattern. Neutralization of the chitosan forms a gel-like precipitate, and the hydrostatic pressure in the sodium hydroxide (NaOH) solution keeps the cuboid scaffold in shape. Comparison of the freeze-dried scaffold to the wet one showed linear and volumetric shrinkage of about 31% and 62%, respectively. A good attachment between layers allowed the chitosan matrix to form a fully interconnected channel architecture. Results of in vitro cell culture studies revealed the scaffold biocompatibility. The results of this preliminary study using the rapid prototyping robotic dispensing (RPBOD) system demonstrated its potential in fabricating three-dimensional (3D) scaffolds with regular and reproducible macropore architecture.     

Facile solution synthesis and characterization of CaCO3 microspheres with urchin-shaped structure

Urchin-shaped CaCO₃ microspheres were successfully synthesized by a facile low-temperature solution-phase method. The product was characterized with X-ray diffraction (XRD), Raman spectra and high resolution transmission electron microscopy (HRTEM). The characterization results revealed that the structure of the urchin-shaped CaCO₃ was grown along the (104) crystal plane with a calcite structure. Furthermore, the valence states and the surface chemical compositions of CaCO₃ were further identified by X-ray photoelectron spectroscopy (XPS). This study provides a simple method to prepare urchin-shaped CaCO₃ microspheres in large scale, which broads their practical applications.     

Gold nanoparticle-mediated fluorescence enhancement by two-photon polymerized 3D microstructures

Fluorescence enhancement achieved by functionalized microstructures made by two-photon polymerization (TPP) is reported for the first time. Microstructures of various shapes made of SU-8 photoresist were prepared and coated with gold nanoparticles (NP) of 80 nm. Localized fluorescence enhancement was demonstrated by microstructures equipped with tips of sub-micron dimensions. The enhancement was realized by positioning the NP-coated structures over fluorescent protein layers. Two fluorophores with their absorption in the red and in the green region of the VIS spectrum were used. Laser scanning confocal microscopy was used to quantify the enhancement. The enhancement factor was as high as 6 in areas of several square-micrometers and more than 3 in the case of local enhancement, comparable with literature values for similar nanoparticles. The structured pattern of the observed fluorescence intensity indicates a classic enhancement mechanism realized by standing waves over reflecting surfaces. With further development mobile microtools made by TPP and functionalized by metal NPs can be actuated by optical tweezers and position to any fluorescent micro-object, such as single cells to realize localized, targeted fluorescence enhancement.     

Effects of heat treatments on the microstructures and mechanical properties of Mg–3Nd–0.2Zn–0.4Zr (wt.%) alloy

Microstructure and mechanical properties of as-cast and different heat treated Mg–3Nd–0.2Zn–0.4Zr (wt.%) (NZ30K) alloys were investigated. The as-cast alloy was comprised of magnesium matrix and Mg₁₂Nd eutectic compounds. After solution treatment at 540 °C for 6 h, the eutectic compounds dissolved into the matrix and small Zr-containing particles precipitated at grain interiors. Further aging at low temperatures led to plate-shaped metastable precipitates, which strengthened the alloy. Peak-aged at 200 °C for 10–16 h, fine β″ particles with DO₁₉ structure was the dominant strengthening phase. The alloy had ultimate tensile strength (UTS) and elongation of 300–305 MPa and 11%, respectively. Aged at 250 °C for 10 h, coarse β′ particles with fcc structure was the dominant strengthening phase. The alloy showed UTS and elongation of 265 MPa and 20%, respectively. Yield strengths (YS) of these two aged conditions were in the same level, about 140 MPa. Precipitation strengthening was the largest contributor (about 60%) to the strength in these two aged conditions. The hardness of aged NZ30K alloy seemed to correspond to UTS not YS.     

Characterization of tensile properties and microstructures in directionally solidified Al–Si alloys using linear roughness index

The tensile properties in directionally solidified Al–Si alloys and in gravity die casting of the same composition are presented. Examples of relating both the tensile and the microstructural properties of these alloys with the fracture roughness index are indicated. The roughness index was measured on vertical sections cut through the tensile fracture surface. The tensile properties examined were the yield and ultimate tensile strengths, strain at fracture, and Young's modulus. The analyzed microstructural features were porosity, dendrite area fraction, secondary dendrite arm spacing, and Si particle spacing. In almost all cases an unambiguous correlation was found between the roughness index and the tensile or the microstructural properties. A marked improvement in ductility was observed for directionally solidified samples over their gravity die casting processed counterparts. The roughness index diminished going from die cast to direction solidification, and this is likely accompanied by change in the fracture mechanisms.     

Earthquake‐induced hydrodynamic pressures on a 3D rigid dam–reservoir system using DRBEM and a radiation matrix

The dual reciprocity method is applied to determine the hydrodynamic pressure distribution in a three‐dimensional dam–reservoir system subjected to earthquake excitation. The reservoir domain is idealized as a finite region of irregular geometry adjacent to the dam followed by an infinite domain of uniform cross‐section in the upstream direction. The reservoir hydrodynamic pressure response is governed by the Helmholtz equation subject to free surface, dam–reservoir interface, absorbing bottom/banks and radiation boundary conditions. A three‐dimensional (3D) dual reciprocity model is developed to determine the hydrodynamic pressure in the finite reservoir domain. A radiation matrix is developed and introduced to relate the hydrodynamic pressure and its normal derivative on the interface between the finite and infinite domains. The three‐dimensional radiation model used is developed by applying a two‐dimensional dual reciprocity formulation along the interface of the finite and infinite reservoirs together with a continuum solution in the upstream direction of the infinite domain. The model is compared for the hydrodynamic response of a three‐dimensional rectangular reservoir and found to be in excellent agreement with results obtained from a model based on the analytical formulation existing in the literature. Copyright © 2003 John Wiley & Sons, Ltd.     

Synthesis of MgCO3 microcrystals at 160 °C starting from various magnesium sources

Rhombohedra microcrystallites of magnesium carbonate (MgCO₃) have been hydrothermally synthesized by urea and magnesium sources (Mg, MgCl₂, Mg(OH)₂, MgSO₄, and MgO) at 160 °C. The experimental parameters such as reaction temperature, time, and reactants were researched to investigate the reaction mechanism. The product synthesized by Mg powder was about 10 μm, and the diameters of the products synthesized by Mg(OH)₂ and MgSO₄ were about 1-2 μm. The product could transfer from the mixture of MgCO₃, Mg(OH)₂, Mg₅(CO₃)₄(OH)₂·4H₂O to pure MgCO₃ by increasing reaction time or raising reaction temperature, which indicates that Mg₅(CO₃)₄(OH)₂·4H₂O was just the intermediate product, and MgCO₃ was the final product.     

Facile synthesis and characterization of CdTe quantum dots–polystyrene fluorescent composite nanospheres

A novel method for the synthesis of CdTe quantum dots–polystyrene (CdTe/PS) composite nanospheres was developed by a two-step route. The as-prepared composite nanospheres were characterized by high-resolution transmission electron microscopy (HRTEM), X-ray powder diffraction (XRD), fluorescent microscope imaging and fluorescence spectrometer. The results showed that the composite nanospheres were spherical and relatively uniform with a diameter of 120 nm. Compared with previous methods, the proposed method was simple, efficient and inexpensive, moreover the composite nanospheres exhibited favorable water-dispersible, stable, size-tunable and fluorescent properties.     

Permanent 3D microstructures in a polymeric host created using holographic optical tweezers

Abstract

Permanent 3D microstructures are created within a gel using holographic optical tweezers. The micron-sized particles are arranged in a precise geometry within an appropriate liquid, and become fixed in position upon polymerisation of the surrounding media. The flexibility of the holographic approach enables any structural arrangement to be produced, dependent upon application. We demonstrate the technique by creating structures within a biocompatible host, enabling future applications in biotechnology.     

Comparative characterization to the three kinds of solid-shape FeS

The microstructures of three kinds of the synthetical solid FeS, acting as the solid lubricant, which includes FeS bulk, FeS particles and FeS powder, were studied in this article. Their surface morphologies were investigated with SEM, and XRD was used to study the phase structures, TEM was utilized to observe the micro-morphologies and analyze the microstructures. The results showed that the crystals of solid FeS were composed of many hexagonal single crystals and multi-crystals, with two kinds of crystalline structure, hexagonal structure and face-centered cubic structure. The surface of FeS was in the shape of a floccule except for a little granular figure. The micro-morphology of FeS presented irregular flakes which can be seen from TEM pictures, the size of grains ranging between 0.1 μm and 3 μm.     

Microwave synthesis and characterization of ZnO with various morphologies

ZnO micro- and nanostructures with a variety of morphologies have been synthesized using Zn(NO₃)₂·6H₂O and pyridine by a microwave-assisted aqueous solution method at 90 °C for 10 min. The pyridine has a significant influence on the morphology of ZnO. Various morphologies of ZnO (hexagonal columns, linked hexagonal needles, hollow structures, and hexagonal nanorings) were obtained by adjusting the concentration of pyridine. The effect of the type of other alkaline additive (aniline and triethanolamine) on the morphology of ZnO was also investigated. The products were characterized by X-ray powder diffraction (XRD), scanning electron microscopy (SEM), transmission electron microscopy (TEM) and high-resolution transmission electron microscopy (HRTEM).     

Evaluation of microstructures and mechanical properties of diamond like carbon films deposited by filtered cathodic arc plasma

Diamond-like carbon (DLC) films were deposited by a cathodic arc plasma evaporation (CAPD) process, using a mechanical shield filter combined with a magnetic filter with enhanced arc structure at substrate-bias voltage ranging from − 50 to − 300 V. The film characteristics were investigated using Raman spectroscopy, X-ray photoelectron spectroscopy (XPS), atomic force microscopy (AFM) and high-resolution transmission electron microscopy (HRTEM). The mechanical properties were investigated by using a nanoindentation tester, scratch test and ball on disc wear test. The Raman spectra of the films showed that the wavenumber ranging from 900 to 1800 cm^− 1 could be deconvoluted into 1140 cm^− 1, D band and G band. The bias caused a significant effect on the sp³ content which was increased with the decreasing of I_D/I_G ratio. The XPS spectra data of the films which were etched by H⁺ plasma indicated the sp³ content are higher than those of the as-deposited DLC films. This implied that there is a sp²-rich layer present on the surface of the as-deposited DLC films. The nanoindentation hardness increased as the maximum load increased. A 380 nm thick and well adhered DLC film was successfully deposited on WC-Co substrate above a Ti interlayer. The adhesion critical load of the DLC films was about 33 N. The results of the wear tests demonstrated that the friction coefficient of the DLC films was between 0.12 and 0.2.     

Formation of intermetallic compounds in the Cr–Al–Si ternary system

The microstructure and composition of binary and ternary intermetallics have been studied in ternary diffusion couples of Cr and an Al–Si eutectic alloy. The ternary intermetallic always formed in the liquid part of the diffusion couple as a dendritic structure. Two intermetallics compounds, CrSi₂ and Cr₅Si₃, of the Cr–Si binary system have been observed. The CrSi₂ intermetallic has a high solubility of up to 20 at.% Al and forms as faceted plates. A number of intermetallics, namely, CrAl₇, Cr₂Al₁₁, CrAl₄, Cr₄Al₉, Cr₅Al₈ and Cr₂Al, of the Cr–Al system have been observed. The solubility of Si varies from as low as 0.8 at.% in Cr₂Al to as high as 9 at.% in Cr₄Al₉. A schematic of the reaction scheme of the Cr–Al–Si system is presented. This has been based on the observed microstructure and composition of phases.     

In situ synthesis mechanism and characterization of ZrB2–ZrC–SiC ultra high-temperature ceramics

The synthesis route, microstructure and properties of ZrB₂–ZrC–SiC composites prepared from a mixture of Zr–B₄C–Si powders by in situ reactive synthesis were investigated. The reactive path and synthesized mechanism of ZrB₂–ZrC–SiC composite were studied through series of pressureless heat treatments ranging from 800 °C to 1700 °C in argon. The in situ ZrB₂–ZrC–SiC composites were fabricated under different synthesis processing. The one with 88.4% relative density performed poorly in mechanical properties due to the occurring of self-propagating high-temperature synthesis (SHS). The fully dense ZrB₂–ZrC–SiC composite was obtained under the optimized synthesis processing without SHS reactions. Its Vickers hardness, flexural strength and fracture toughness were 20.22 ± 0.56 GPa, 526 ± 9 MPa and 6.70 ± 0.20 MPa m^1/2, respectively.     

Facile synthesis via a solvothermal route and characterization of Mg–Al layered double hydroxide (LDH) 3D micro–nano structures

Different Mg–Al layered double hydroxide (LDH) 3D micro–nano structures had been synthesized via a facile solvothermal method. By carefully controlling the fundamental experimental parameters, the morphologies of hexagonal nanoplates, rose-like micro–nano structures, chrysanthemum-like micro–nano structures, and spherical micro–nano structures have been efficiently obtained, respectively. And these micro–nano structures are formed from self-assembly of nanoplates in a spontaneous process in solvothermal system. It is also found that the concentration of the reactant and the dosage of CTAB have significant effects on the morphology of the products. The FT-IR spectra and thermal stability of the micro–nano structures were explored.