|
Wednesday 10 November 1999, Kelvin Conference Centre, University of Glasgow.
|
|
Adam Curtis,Institute of Biomedical and Life Sciences, University of Glasgow. The
contact between cell and cell and between cell and a material
substrate is the region in which adhesions form and are maintained.
Though very useful information can be obtained from TEM direct
observation of this zone in life is difficult. I shall describe
three optical methods, which can be used to examine this region
when the cells are grown on surfaces whose refractive index differs
appreciably from that of the medium or when appropriate fluorescent
labelling can be used. These methods are Interference Reflection
Microscopy (IRM), Surface Plasmon Resonance Microscopy (SPRM)
and Fluorescence Energy Transfer Microscopy (FETM). Each of these
methods has a resolution in the z-axis of at least 10nm and FETM
in fact yields no signal when the cell is more than 6nm from
the surface. IRM is also a very sensitive method for detecting
surface contamination. I shall describe the methods, how they
work and how they may be set up and something of the information
they give on this nearly two-dimensional world under the cell. Mazz Marry1,
I. Max Huxam2, Laurence Tetley2 and Michael C. Jarvis1, The cell wall of higher plants is composed of at least two major structurally independent but interacting networks. The cellulose-hemicellulose network (about 50% of the wall mass), is embedded in a second network of matrix pectic polysaccharides (about 30% of the total mass). There are many types of cross-links involved in holding carbohydrate moieties within the plant cell wall, all of which directly influence cell-cell adhesion and the mechanical strength of the cell wall. Various techniques can be used to analyse the nature and distribution of the carbohydrate polymers present in plant cell walls. Three such techniques, Fourier Transform Infra-red (FTIR) microspectroscopy, immunogold labelling by TEM and Electron Energy Loss Spectroscopy (EELS) were used to investigate the cell walls from mature sugar beet (Beta vulgaris L. Aztec). Unextracted cell walls and the wall residue remaining following four sequential extractions with a calcium chelator (imidazole) and two additional extractions with sodium carbonate (to remove ester bonds) were studied. Unlike
most dicots, sugar beet have significant levels of ferulic acid
in their primary walls. These form ester cross-links, which may
strengthen the cell wall. FTIR analysis indicates that the wall
residue following imidazole extraction contains a high proportion
of both saturated and unsaturated esters, which are removed from
the wall by the additional treatment of sodium carbonate. Immuno-gold
labelling has provided information on the distribution of both
unesterified and methyl-esterified pectic epitopes within the
cell wall residue. Successive imidazole extracts removed unesterified
pectic epitopes throughout the cell wall, but not from the middle
lamella. Elemental maps from the unextracted wall material clearly
show calcium ions throughout the wall, with a greater concentration
in the middle lamella. It has been known for some time that,
in mature tissues, the middle lamella glues together the walls
of contiguous cells and forms a separate domain within the wall
which mainly consists of calcium-cross-linked pectic polysaccharides,
polymers which are rich in galacturonic acid. The EELS-imaging
data of the wall residue following imidazole extractions indicate
that the unesterified pectic epitopes still present within the
middle lamella are not cross-linked by calcium ions. Alan Craven, Department of Physics and Astronomy, University of Glasgow, Glasgow.G12 8QQ. Electrons have a number of advantages when it comes to using them to study materials. On the optical side, it is relatively easy to make high brightness sources of highly monochromatic electrons in the laboratory. These electrons can be focused with simple magnetic lenses, allowing the formation of either a high-resolution image of the electrons leaving a thin specimen, as in the transmission electron microscope, or a very small electron probe, as in the case of the scanning electron microscope. On the interaction side, the electrons are scattered quite strongly both elastically and inelastically, leading to a wide range of signals that can provide information about the materials being studied. The skill of the electron microscopist comes in making use of these signals to provide the information sought about the material. The talk will show how such signals can be used to study things like paints, hard wear-resistant coatings, pipeline steels and magnetic devices.
F. T. Docherty*§,
A. J. Craven§ and D. W. McComb*, Electron energy loss spectroscopy (EELS) provides information about the energy loss suffered by primary electrons when they interact with a specimen. The resulting energy loss spectrum contains edges due to the excitation of core electrons to unoccupied states in the conduction band. These edges exhibit fine structure, which is dependent on the local chemical and structural environment of the absorbing atom. The understanding of this technique can be improved by examining data collected from a series of compounds with the same crystal structures, to investigate the effect on the fine structure of changing the element in a specific crystallographic site. Spinels
are a suitable class of compounds for a study of this type. The
study of a series of chromium spinels, [A]tet[Cr2]octO4, (A=Mg2+,
Co2+, Ni2+, Cu2+, Zn2+) will determine the effect on the Cr and
O edges of occupancy of the d-orbitals on the neighbouring sites.
By correlating the experimental data with electronic structure
calculations, we aim to develop an interpretative framework,
which can be used to determine the chemistry of more complex
materials, improving the fundamental understanding of ELNES. Sally E. Solomon, Poultry Research Unit, Department of Preclinical Veterinary Studies, University of Glasgow Veterinary School, Bearsden, Glasgow G61 1QH. Technology leads knowledge? and in the case of the avian ovum, suspended in its protective albumen sac and encased in a bioceramic shell, the egg, as we know it, has been the testing ground for both traditional and space age methodology. This paper deals primarily with shell construction and to that end egg white is cited as the foundation layer upon and around which subsequent layers will deposit. Its degree of stiffness is therefore of paramount importance. Low Resolution Nuclear Magnetic Resonance is used to monitor the observed changes in albumen viscosity by measuring its water proton longitudinal relaxation rate. Light, scanning and transmission microscopy have all had a role to play in resolving the morphology of the eggshell proper and scanning microscopy in particular has enabled the characterisation of the twelve major structural variations which occur in the basal layer of the calcitic eggshell and which in turn influence the direction of further crystal growth The Proscan 1000 non-contact surface profiler provides a rapid and objective analysis of the contours of the inner surface of the eggshell. A non-invasive measurement of egg quality is now possible using acoustic resonance (Egg Tapping). The paper will illustrate its use and limitations.
A. Sahal,
M.H.Gladden, D. J. Maxwell and E Jankowska, The modualtory effects of serotonin are facilitatory on both a- and g?motoneurones. In contrast, noradrenaline has facilitatory effects on a?motoneurones but depressive effects on g-motoneurones. Since these differences might be reflected in the coupling between descending noradrenergic and serotoninergic nerve fibres with g-motoneurones the coupling was investigated. Eight g-motoneurones were labelled with rhodamine-dextran injected intracellularly in anaesthetised cats. Serotoninergic and noradrenergic axons were identified with antibodies against dopamine b-hydroxylase and serotonin. Appositions between labelled axons and somata and dendrites were located with three colour confocal laser scanning microscopy by examining series of optical sections gathered at 1mm intervals in 50mm thick sagital sections of the spinal cord. Serotoninergic and noradrenergic varicosities were found in apposition and close to g-motoneurones. Serotoninergic varicosities formed 1.05 ± 0.10 per 100mm2 with the somata and 14.5 ± 1.95, 11.3 ± 1.09 and 7.4 ± 2.29 appositions per mm length of dendrite within 100, 100-300mm and >300mm distances from the soma, respectively. The number of appositions of noradrenergic varicosities were 0.6 ± 0.10 per 100mm 2 with somata and 9.2 ± 1.83, 7.6 ± 1.09 and 3.6 ± 2.37 per 100mm length of the dendrites 100, 100-300 and >300mm from the soma respectively. No major differences were found in the coupling between serotoninergic fibres compared with noradrenergic fibres and g-motoneurones except that the density of serotoninergic appositions was higher. Comparison with data available for coupling between serotoninergic fibres and a-motoneurones suggests that it is similar for a- and g-motoneurones. Comparison of the numbers of varicosities in apposition to those in the immediate surround (in a 5mm shell) suggests that the varicosities are preferentially located on the surface of the g-motoneurones.
ACQUISITION OF DIGITAL DATA IN THE MODERN TEM Timon F.
Fliervoet, Max T. Otten, Wim M. Busing and Marc H. Storms, FEI
I Philips Electron Optics, Eindhoven, The Netherlands. A thorough characterisation of a material is the start of a fundamental understanding of the material's properties. Morphology, crystal structure, chemical composition, interface structure, surfaces and defects all have their influence on the properties of materials. In (advanced) materials science and design there is an increasing need for the determination of the chemical and electronic structure of materials. As microstructures become more and more complex, the dimensions to be analysed are becoming smaller. Transmission Electron Microscopy (TEM) has proven to be a very powerful technique for studying a range of materials down to sub-nanometer levels. It generates a wide range of signals, carrying different types of valuable information. TEM also contributed significantly in finding answers to fundamental questions about the functionality of ongoing physiological and pathological processes. However, in the past TEMs, equipped with many detectors, were surrounded by a single proliferation with PC's, monitors and mice. These add-on monitors and computers often caused stray-fields and interferences, and require too much room. Furthermore, it was difficult to operate the different computers with different operating systems. Most important, the interaction between microscopy and the data was lost. The Tecnai series of TEM has been especially designed to acquire and process these digital signals efficiently and simultaneously. All the detectors - including STEM, TV and CCD cameras, EDX and EELS detectors - are fully embedded into one Tecnai operating system, with one monitor, mouse and keyboard. Tecnai, the modem TEM, makes microscopy easier then ever.
W. L. Maxwell and M. Nielson, Laboratory of Human Anatomy, University of Glasgow, Glasgow G12 8QQ. Axons
within the white matter of the brain are exposed to transient
tensile strain during diffuse axonal injury (DAI) which is the
commonest cause of death in children, and adults under the age
of 40. A great deal of attention in a number of experimental
laboratories over the last 15 years has been focused upon the
characterisation of axonal pathology with the aim of defining
the pathological mechanisms operating in such models of DAI.
It is clear that axons are not fragmented at the time of injury
but rather enter a so-called "pathological cascade"
of events which lead to axotomy over hours and days after injury.
A Kontron Elektronik-Vidas Image Analysis has been used to develop
a program for the quantitative analysis of responses by components
of the axonal cytoskeleton, microtubules and neurofilaments,
to the application of transient mechanical strain. |