On the preparation, microscopic investigation and application of ISCOMs

ISCOM matrices constitute colloidal structures formed from Quillaja saponins, cholesterol and phospholipid. Addition of protein antigens to these matrices leads to the formation of ISCOMs. In this review we report on microscopic investigations of ISCOM matrices and ISCOMs as well as related colloidal structures, such as helices, worm-like micelles, ring-like micelles, and lamellae structures. We briefly outline the immunologic basis for the use of ISCOMs as vaccine delivery systems, and describe the various methods to form ISCOMs. Negative staining transmission electron micrographs of all colloidal structures are presented and described. On the basis of our microscopic investigations, different formation mechanisms of ISCOMS are discussed.     

Continuous and time-shared multiple optical tweezers for the study of single motor proteins

We present a comparison between continuous (CW) and time-shared (TS) multiple optical tweezers applied to the study of the interaction between a single motor protein (myosin) with its track (an actin filament). In the experimental assay, named “three-bead assay”, a single actin filament is stretched between two beads trapped in a CW or in a TS double trap. The actin filament is presented to a single myosin molecule lying on a third bead attached to the coverslide. The CW double trap is obtained by splitting a single laser source into two orthogonally polarized beams, while the TS one is obtained by rapidly scanning a single laser beam with acusto-optic modulators. When using the CW traps, position detection of the left or right bead is obtained by means of a single quadrant detector photodiode (QDP) placed in the back focal plane of the condenser and selected with a polarizer; when using the TS traps, the position of multiple traps with the same QDP can be collected independently using triggered and synchronized generation and acquisition. The two techniques are thoroughly analysed and compared, evidencing advantages and disadvantages of each one.     

Ultrastructural examination of dentin using focused ion-beam cross-sectioning and transmission electron microscopy

Focused ion-beam (FIB) milling is a commonly used technique for transmission electron microscopy (TEM) sample preparation of inorganic materials. In this study, we seek to evaluate the FIB as a TEM preparation tool for human dentin. Two particular problems involving dentin, a structural analog of bone that makes up the bulk of the human tooth, are examined. Firstly, the process of aging is studied through an investigation of the mineralization in ‘transparent’ dentin, which is formed naturally due to the filling up of dentinal tubules with large mineral crystals. Next, the process of fracture is examined to evaluate incipient events that occur at the collagen fiber level. For both these cases, FIB-milling was able to generate high-quality specimens that could be used for subsequent TEM examination. The changes in the mineralization suggested a simple mechanism of mineral ‘dissolution and reprecipitation’, while examination of the collagen revealed incipient damage in the form of voids within the collagen fibers. These studies help shed light on the process of aging and fracture of mineralized tissues and are useful steps in developing a framework for understanding such processes.     

明永乐青花瓷片的显微拉曼光谱分析

利用显微拉曼光谱对明永乐青花瓷釉及釉中的结晶物进行了分析,釉的拉曼谱中Si-O弯曲振动与伸缩振动峰的面积比(Ip=A500/A1000)与陶瓷的烧成温度相关连。同时在釉中还发现有Fe3O4、α-Fe2O3、Co3O4、MnO、Mn2O3、硅酸钙、磷酸钙等结晶物。研究结果表明,显微拉曼光谱是一种简便、可靠的无损检测手段,对古陶瓷的鉴定与分析有重要的应用意义。     

The imaging of nano and micro globules of short linear thermo responsive polyacrylamides formed above the lower critical solution temperature

An imaging method has been developed to examine thermo responsive polymer coagulates by optical and electron microscopy. Poly-N-isopropylacrylamide (PNIPAM), poly-N-dimethylacrylamide (PDMAM) and a 1:1 PNIPAM-PDMAM copolymer were encapsulated in a gelatin matrix as coagulates above the lower critical solution temperature (LCST), and subsequently examined by optical and electron microscopy. The linear macromolecules PNIPAM and PDMAM were synthesized by chain transfer polymerization with mercaptopropionic acid (3-MPA) as chain transfer reagent. The resulting polymers have an average molar mass of ∼1800 g/mol and low polydispersity. The LCST of thermo responsive polymers is defined in pure water but can also be stimulated at lower than the phase transition temperature employing electrolytes containing inorganic salts such as (NH₄)₂SO₄. Under such conditions the polymers show the typical thermo responsive phase transfer property in form of a visible clouding point. Gelatin was used to maintain this biphasic state by slowly adding water-softened gelatin sheets at a temperature above the LCST, followed by cooling to 3 °C in order to induce gelation. Examination of the gelatin-coagulate matrices by optical and electronic microscopy showed that PNIPAM and its copolymer (PNIPAM/PDMAM 1:1) are entrapped as globular spheres and clusters of spheres. In comparison pure PDMAM, even if it shows a clouding point, does not form typical LCST coagulates. With PNIPAM and the copolymer, micro globule formation is also possible with slow gelatin formation, without first provoking an LCST. In this particular case, the phase transition, or entropic demixing of the polymers respectively, are induced in this case by water absorption of the gelatin matrix.     

Effect of potassium permanganate on morphological,structural and electro-optical properties of graphene oxide thin films

This work investigated the effect of Potassium Permanganate (KMnO₄) on graphene oxide (GO) properties, especially on electrical properties. The GO thin films were deposited on a glass substrate using drop casting technique and were analysed by using various type of spectroscopy (e.g. Scanning Electron Microscopy (SEM), Ultra- Violet Visible (UV–VIS), Fourier Transform Infrared (FTIR), X-Ray Diffraction (XRD), optical band gap, Raman Spectroscopy). Furthermore, the electrical experiments were carried out by using current–voltage (I-V) characteristic. The GO thin film with 4.5 g of KMnO₄ resulted in higher conductivity which is 3.11 × 10^?4 S/cm while GO with 2.5 g and 3.5 g of KMnO₄ achieve 2.47 × 10^?9 S/cm and 1.07 × 10^?7 S/cm, respectively. This further affects the morphological (SEM), optical (band gap, UV–Vis, FTIR, and Raman), and crystalline structural (XRD) properties of the GO thin films. The morphological, elemental, optical, and structural data confirmed that the properties of GO is affected by different amount of KMnO₄ oxidizing agent, which revealed that GO can potentially be implemented for electrical and electronic devices.     

Instrument for Hadamard transform three-dimensional fluorescence microscope image analysis

An instrument combining fluorescence microscopy with Hadamard transform multiplexed imaging was designed by which a three-dimensional Hadamard transform fluorescence microscopic cell image was obtained. The image can provide useful information including, simultaneously, the apparent dimensions and the shape of the analytical sample, the content and the distribution of some species in it.     

Model films of cellulose: I. Method development and initial results

This report presents a new method for the preparation of thin cellulosefilms. NMMO (N-methylmorpholine-N-oxide) was used to dissolve cellulose andaddition of DMSO (dimethyl sulfoxide) was used to control viscosity of thecellulose solution. A thin layer of the cellulose solution is spin-coated ontoasilicon oxide wafer and the cellulose is precipitated in deionised water. Thecellulose film is anchored onto the silicon oxide wafer by a saturated polymerlayer. Among many different polymers tested, PVAm (polyvinylamine) and G-PAM(glyoxalated-polyacrylamide) worked well. The preparation of cellulose modelfilms described in this paper resulted in films with thicknesses in the range20–270 nm and the thickness can be controlled by alteringtheconcentration of cellulose solution by addition of different amounts of DMSO.The films were cleaned in deionised water and were found to be free fromsolvents by ESCA analysis and contact angle measurements. The molecular weightdistribution of the cellulose surface material shows that there is only minorbreakdown of the cellulose chains, mainly by cleavage of the longest molecularmass fraction and without creation of low molecular mass oligomers of glucose.     

Supercritical fluid chromatography/Fourier transform infrared microspectrometry

The feasibility of supercritical fluid chromatography/Fourier transform-infrared (SFC/FT-IR) microspectrometry is presented. In this approach to SFC/FT-IR, the chromatographic eluates are aspirated from the restrictor directly onto the surface of a moving window which then passes into the beam focus of an infrared microscope. Because the mobile phase is gaseous at ambient conditions, elimination of the mobile phase is easily accomplished. Detection limits in all interfaces between a chromatograph and an FT-IR spectrometer in which the mobile phase is eliminated are determined in large part by the area over which the sample is deposited. We have shown that SFC eluates can be condensed at ambient temperature into spots of 100 to 200μm in diameter. The microscope interface therefore serves to increase the sensitivity of the SFC/FT-IR measurements of these spots and detection limits in the low nanogram range are possible. Preliminary results obtained before any real attempts were made to optimize the deposition process indicate that identifiable spectra can be obtained in real time at the 50 ng level for chromatographic separations performed with a 100μm i.d. wall coated open tubular column. Reproducible reconstructed chromatograms are obtained as each deposited eluate travels through the beam focus of the microscope. The concentration profile of deposited peaks was determined by IR measurements performed at 50 μm increments over the width of the peak to ascertain the deposition size. The results described in this paper, while not yet optimized, indicate the great potential of SFC/FT-IR microspectrometry.