Freeze shattering: a simple and effective method for permeabilizing higher plant cell walls

This article describes a practical technique for permeabilization of higher plant cell walls, which is usually one of the first steps required for immunolocalization of cellular components (and other cytological methods) in plant cell studies. Our strategy involves shattering the walls of cells while the tissues are frozen in liquid nitrogen. It replaces the use of wall degrading enzymes or the need to employ laborious sectioning or other mechanical means for providing access of probes to cells. Freeze-shattering retains the integrity of whole tissues and cells surprisingly well and thus is especially useful when used in conjunction with confocal laser scanning microscopy for recording the three-dimensional arrangement of cytoskeletal elements in relation to cell shape. In this article, we demonstrate the effectiveness of this technique for anti-tubulin and anti-actin immunofluorescence and for rhodamine phalloidin labelling of the cytoskeleton in various higher plant tissues including onion root tip and bulb scale epidermis, Tradescantia stamen hairs and Arabidopsis leaf epidermis and mesophyll cells.     

A new technique for making plant epidermal imprints using various domestic adhesives

A simple, quick and easy method for making leaf surface imprints for microscopic studies by using cheap and easily available non-toxic domestic adhesives is described. The most satisfactory results were obtained, in order of preference, by using ‘Quickfix’, ‘Stickfast’, ‘Fevicol’, ‘D.P.X. (mountant)’, ‘Camel paste’, ‘Cello-tape’ and ‘Araldite’. These were used on the epidermis of vegetative and floral organs of a number of plants (living and dried). A semi-transparent film was allowed to dry, which was then stripped off and mounted on a clean slide. This film records very fine structural details which are evident from the photomicrographs presented. With the help of this technique, the form, distribution and orientation of stomata, size of guard cells and their movement, size of stomatal pores, stomatal development, cell-wall nature, stomatal frequency and indices and venation pattern can easily be studied. Repeated imprints can be made from the same surface of an organ. This method should be of great use to plant morphologists and anatomists.     

Microtubule and cellulose microfibril orientation during plant cell and organ growth

In this review, I ask the question of what is the relationship between growth and the orientations of microtubules and cellulose microfibrils in plant cells. This should be a relatively simple question to answer considering that text books commonly describe microtubules and cellulose microfibrils as hoops that drive expansion perpendicular to their orientation. However, recent live imaging techniques, which allow microtubules and cellulose synthase dynamics to be imaged simultaneously with cell elongation, show that cells can elongate with nonperpendicular microtubule arrays. In this review, I look at the significance of these different microtubule arrangements for growth and cell wall architecture and how these resultant walls differ from those derived from perpendicular arrays. I also discuss how these divergent arrays in stems may be important for coordinating growth between the different cell layers. This role reveals some general features of microtubule alignment that can be used to predict the growth status of organs. In conclusion, nonperpendicular arrays demonstrate alternative ways of cell elongation that do not require hooped arrays of microtubules and cellulose microfibrils. Such nonperpendicular arrays may be required for optimal growth and strengthening of tissues.     

Study of deposit and friction films of overbased calcium sulphonate by PM-IRRAS spectroscopy

This paper deals with the study of deposit and friction films of overbased calcium sulphonate diluted in mineral oil, on metal surfaces. The technique used for this work is Infrared Reflection Absorption Spectroscopy by Fourier Transform and Polarisation Modulation (PM-IRRAS or PM FTIR). The spectra obtained from this method contain only information on the absorption occurring in the immediate neighbourhood (⩽40 nm) of the metallic substrate and allow the average orientation for molecules adsorbed on metals to be deduced. This study shows that the adsorption of overbased calcium sulphonate on a steel surface leads to a preferential orientation of the sulphonate chains perpendicular to the surface and to a preferential orientation of the carbonate, the c axis being perpendicular to the surface. During friction, the sulphonate chains are ejected from the contact zone. We observed that the boundary film consists mainly of calcium carbonate, which crystallises into calcite.     

Infra-red microspectroscopy of hydrated biological systems: design and construction of a new cell with atmospheric control for the study of plant cell walls

A new hydration cell has been constructed that allows wet biological samples, or samples of controlled moisture content, to be analysed in situ using infra-red microspectroscopy. The cell has been used to show that there are minor spectral changes associated with the hydration of pectin and tomato pericarp cell walls and slightly more significant changes in onion and carrot epidermal walls. The cell was also used to show that molecular orientations of polymers, previously observed in dry cell walls, were also to be seen in hydrated walls. For cell walls of onion and carrot epidermis, it was shown that the orientations of cell wall polymers are not affected by hydration. Furthermore, the polymer orientations in cell walls of fully elongated onion epidermal cells are different from those of elongating carrot epidermis. By using the hydration cell, it is now possible to investigate both fresh samples and wet systems routinely. The applications of this to the study of biological materials with infra-red microspectroscopy are discussed.     

An orientation‐independent DIC microscope allows high resolution imaging of epithelial cell migration and wound healing in a cnidarian model

Epithelial cell dynamics can be difficult to study in intact animals or tissues. Here we use the medusa form of the hydrozoan Clytia hemisphaerica, which is covered with a monolayer of epithelial cells, to test the efficacy of an orientation‐independent differential interference contrast microscope for in vivo imaging of wound healing. Orientation‐independent differential interference contrast provides an unprecedented resolution phase image of epithelial cells closing a wound in a live, nontransgenic animal model. In particular, the orientation‐independent differential interference contrast microscope equipped with a 40x/0.75NA objective lens and using the illumination light with wavelength 546 nm demonstrated a resolution of 460 nm. The repair of individual cells, the adhesion of cells to close a gap, and the concomitant contraction of these cells during closure is clearly visualized.     

The role of the plant cytoskeleton in the interaction between legumes and rhizobia

The study of the symbiotic interaction between rhizobia and legumes represents a major theme in plant biology. This interaction results in the formation of nodules, root organs in which the bacteria reduce atmospheric nitrogen into ammonia, which can subsequently be utilized by the plant. The execution of the different developmental stages observed during nodule ontogenesis involves many cellular processes with significant roles for the plant cytoskeleton. A challenging question in cell biology is how the cytoskeleton organizes itself into the dynamic arrays required for cell differentiation and functioning. Nodulation is, particularly, well qualified as an experimental system for cytoskeleton research because an early essential step of the plant/microbe interaction takes place in surface-exposed root hairs, well suited for cell biological in vivo experimentation. Moreover, the changes in the organization of the cytoskeleton can be elicited by a well-defined molecule, the Nod factor, or by bacterial inoculation, thus providing the researcher with the possibility of controlling the cytoskeletal changes in target cells. In addition, the well-known cytology of the symbiotic interaction facilitates the correlation between the changes in the organization of the plant cytoskeleton with both histological and cellular changes. In this review, the current knowledge on the role of the plant cytoskeleton during nodulation is summarized, with emphasis on the interaction between Medicago truncatula and Sinorhizobium meliloti .     

A ranking method based on DEA and PROMETHEE II (a rank based on DEA & PR.II)

In this paper we present an integrated Data Envelopment Analysis-Multiple Criteria Decision Aid (DEA-MCDA) model which can be applied to increase the discrimination power of DEA. The aim is to restrict weight values of a DEA model by using tools from MCDA. This model leads to more reasonable inputs/outputs weights while in classic DEA models, some inputs/outputs may be characterized by very low or high weight values. To achieve this goal we use the stability intervals based on PROMETHEE II (Preference Ranking Organization METHod for Enrichment of Evaluations) as weight constraints in DEA. Furthermore, the unicriterion net flow scores matrix is used instead of the initial evaluation matrix. By doing so, we already integrate preferential information in the DEA process. By construction, the best results are compatible with the PROMETHEE II ranking. Additional comparisons with the outputs of other decision making techniques are provided based on two examples.     

The blood-testis barrier and Sertoli cell junctions: structural considerations.

In this review, a few well-established axioms have been challenged while others were viewed from a new perspective. The extensive literature on the blood-testis barrier has been scrutinized to help probe its mechanics and hopefully to promote understanding of the constant adaptation of the barrier function to germ cell development. Our principal conclusions are as follows: (1) Although the barrier zonule is topographically located at the base of the seminiferous epithelium it actually encircles the apex of the Sertoli cell. Consequently the long irregular processes specialized in holding and shaping the developing germ cells should be considered as apical appendages analogous to microvilli. (2) The development of the barrier zonule does not coincide with the appearance of a particular class of germ cells. (3) The barrier compartmentalizes the epithelium into only two cellular compartments: basal and lumenal. (4) Although the blood-testis barrier does sequester germ cells usually considered antigenic, immunoregulator factors other than the physical barrier seem to be involved in preventing autoimmune orchitis. (5) Structurally, a Sertoli cell junctional complex is composed of occluding, gap, close, and adhering junctions. The Sertoli cell membrane segments facing germ cells are presumably included in the continuum of the Sertoli cell junctional complex that extends all over the lateral and apical Sertoli cell membranes. (6) The modulation (i.e., formation and dismantling) of the junctions in a baso-apical direction is characteristic of the seminiferous epithelium and may be dictated by germ cell differentiation. The formation of tubulobulbar complexes and the following internalization of junction vesicles conceivably represent sequential steps of a single intricate junction elimination process that involves junction membrane segments from different cell types as part of a continual cell membrane recycling system. (7) The preferential association of junctional particles with one or the other fracture-face reflect a response to various stimuli including seasonal breeding. Changes in the affinity of the particles are generally coincidental with cytoskeletal changes. However, changes in the cytoskeleton are not necessarily accompanied by permeability changes. The number of strands seems to reflect neither the junctional permeability nor the transepithelial resistance. The diverse orientation of the strands seems to be related to the plasticity of the Sertoli cell occluding zonule. (8) Cooperation between all constituents (Sertoli cells, myoid cells, cell substratum, and germ cells) of the epithelium seems essential for the barrier zonule to function in synchrony with the germ cell differentiation.(ABSTRACT TRUNCATED AT 400 WORDS)     

Effects of key operating parameters on the efficiency of two types of pem fuel cell systems (High-Pressure and Low-Pressure operating) for Automotive Applications

The proton exchange membrane (PEM) fuel cell system consisting of stack and balance of plant (BOP) was modeled in a MATLAB/Simulink environment. High-pressure operating (compressor type) and low-pressure operating (air blower type) fuel cell systems were considered, The effects of two main operating parameters (humidity and the pressure of the supplied gas) on the power distribution characteristics of BOP and the net system efficiency of the two systems mentioned above were compared and discussed. The simulation determines an optimum condition regarding parameters such as the cathode air pressure and the relative humidity for maximum net system efficiency for the operating fuel cell systems. This study contributes to get a basic insight into the fuel cell stack and BOP component sizing. Further research using multiobject variable optimization packages and the approach developed by this study can effectively contribute to an operating strategy for the practical use of fuel cell systems for vehicles.     

On suboptimal supervisory control of Petri nets in the presence of uncontrollable transitions via monitor places

Recently, a study has been presented for suboptimal supervisory control of Petri nets via monitor (control) places in the presence of uncontrollable transitions. To enforce a generalized mutual exclusion constraint (GMEC) suboptimally on a plant Petri net with uncontrollable transitions, a set of monitor places has been provided to choose from. There are two main results of this study, the first of which is that “there is not an optimal solution to the problem of computing a monitor-based controller to enforce a GMEC on a plant net in the presence of uncontrollable transitions”. The second result deals with the computation of monitor places for the suboptimal supervisory control in such Petri nets. In this paper, it is shown that there may be an optimal solution to the problem of computing a monitor-based controller to enforce a GMEC on a plant net in the presence of uncontrollable transitions. To do this, an example Petri net is considered and then two different optimal (maximally permissive) solutions are provided for this Petri net.     

DESIGN OF COMBINED CYCLE GENERATION SYSTEM WITH HIGH TEMPERATURE FUEL CELL AND STEAM TURBINE

For environment protection and high efficiency, development of new concept power plant has been required in China. The fuel cell is expected to be used in a power plant as a centralized power station or distributed power plant. It is a chemical power generation device that converts the energy of a chemical reaction directly into electrical energy and not limited by Carnot cycle efficiency. The molten carbonate fuel cell (MCFC) power plant has several attractive features i.e. high efficiency and lower emission of NOx and SOX, A combined cycle generation system with MCFC and steam turbine is designed. Its net electrical efficiency LHV is about 55%.     

Non-zeroth-order Laue zone effects and alchemi

The depth-dependence of fast electron probability density is calculated for the 〈111〉 zone axis orientation of GaAs, and the effects of slight tilt from this orientation. Although Ga and As project onto equivalent sites for beams in the zeroth-order Laue zone, differences in mean probability density on Ga and As occur, due to higher-order Laue zone effects. Whether preferential peaking takes place on Ga or As depends upon which atom is assumed to be in the top surface layer. 64×64 beam multislice calculations are compared with 49-beam Bloch wave computations, in which the fast oscillatory depth-dependence of probability density is not accounted for. These results have implications for the interpretation of dynamical X-ray fluorescence effects, and particularly for site determination under zone axis diffraction conditions.     

A realtime fuzzy Petri net diagnoser for detecting progressive faults in PLC based discrete manufacturing system

In this paper, we explored a realtime fuzzy Petri net approach to diagnose progressive faults in discrete manufacturing systems. Progressive faults are usually caused by deterioration or aging and show stochastic properties. Some researchers have reported how to detect abrupt faults in discrete manufacturing systems using Petri net. However, little research has been conducted on Petri net diagnoser to progressive faults in discrete manufacturing event systems. To tackle this problem, we explored an approach including a realtime Petri net model and a fuzzy Petri net diagnoser to replicate the plant and detect faults in discrete manufacturing systems. The realtime Petri net model monitors events generated from the discrete manufacturing system, also compares the outputs and pre-settings. Once a difference is detected, it will start the fuzzy Petri net diagnoser to locate faults. For the purpose of validation, this approach was implemented with Visual Basic for diagnosing a dual robot arm. Evaluation experiments validated the diagnoser's performance on accuracy and diagnosability. It illustrated that the proposed approach can have a high accuracy rate of 93% and maximum diagnosis delay of eight steps; it proves that the approach has the capability of integrating knowledge and handling uncertainties. It also remedies the nonsynchronization between the diagnoser and the plant. The approach to construct the model and diagnoser is systematic; it has an excellent projection on intermittent fault diagnosis and hybrid systems.     

Micromanipulation by laser microbeam and optical tweezers: from plant cells to single molecules

Complete manipulation by laser light allows precise and gentle treatment of plant cells, subcellular structures, and even individual DNA molecules. Recently, affordable lasers have become available for the construction of microbeams as well as for optical tweezers. This may generate new interest in these tools for plant biologists. Early experiments, reviewed in this journal, showed that laser supported microinjection of material into plant cells or tissues circumvents mechanical problems encountered in microinjection by fragile glass capillaries. Plant protoplasts could be fused with each other when under microscopical observation, and it was no major problem to generate a triple or quadruple fusion product. In the present paper we review experiments where membrane material was prepared from root hair tips and microgravity was simulated in algae. As many plant cells are transparent, it is possible to work inside living, intact cells. New experiments show that it is possible to release by optical micromanipulation, with high spatial resolutions, intracellular calcium from caged compounds and to study calcium oscillations. An example for avian cardiac tissue is given, but the technique is also suitable for plant cell research. As a more technical tool, optical tweezers can be used to spatially fix subcellular structures otherwise moving inside a cell and thus make them available for investigation with a confocal microscope even when the time for image formation is extended (for example at low fluorescence emission). A molecular biological example is the handling of chromosomes and isolated individual DNA molecules by laser microtools. For example, chromosomes can be cut along complex trajectories, not only perpendicular to their long axis. Single DNA molecules are cut by the laser microbeam and, after coupling such a molecule to a polystrene microbead, are handled in complex geometries. Here, the individual DNA molecules are made visible with a conventional fluorescence microscope by fluorescent dyes such as SYBRGreen. The cutting of a single DNA molecule by molecules of the restriction endonuclease EcoRI can be observed directly, i.e. a type of single molecule restriction analysis is possible. Finally, mechanical properties of individual DNA molecules can be observed directly.     

In vivo determination of fibril orientation in plant cell walls with polarization CSLM

Congo Red fluorescence is used to detect cellulose in the wall of plant cells. The orientation of the cellulose fibrils is determined by using polarized light for excitation. The absorption characteristics of Congo Red make this approach a method of choice for applications with any standard confocal scanning laser microscope (CSLM). The semiquantitative character of CSLM observations combined with the non-toxicity of the stain allow a very fast and reliable assessment of cellulose orientation in the wall of living plant cells.     

Quantitative scheme for full-field polarization rotating fluorescence microscopy using a liquid crystal variable retarder

We present a quantitative scheme for full-field polarization rotating fluorescence microscopy. A quarter-wave plate, in combination with a liquid crystal variable retarder, provides a tunable method to rotate polarization states of light prior to its being coupled into a fluorescence microscope. A calibration of the polarization properties of the incident light is performed in order to correct for elliptical polarization states. This calibration allows the response of the sample to linear polarization states of light to be recovered. Three known polarization states of light can be used to determine the average fluorescent dipole orientations in the presence of a spatially varying dc offset or background polarization-invariant fluorescence signal. To demonstrate the capabilities of this device, we measured a series of full-field fluorescence polarization images from fluorescent analogs incorporated in the lipid membrane of Burkitts lymphoma CA46 cells. The fluorescent lipid-like analogs used in this study are molecules that are labeled by either a DiI (1,1(')-Dioctadecyl 3,3,3('),3(')-Tetramethylindocarbocyanine) fluorophore in its head group or a Bodipy (4,4-difluoro-4-bora-3a,4a-diaza-s-indacene) molecule in its acyl chain. A spatially varying contrast in the normalized amplitude was observed on the cell surface, where the orientation of the DiI molecules is tangential to the cell membrane. The internally labeled cellular structures showed zero response to changes in linear polarization, and the net linear polarization amplitude for these regions was zero. This instrument provides a low cost calibrated method that may be coupled to existing fluorescence microscopes to perform investigations of cellular processes that involve a change in molecular orientations.     

Application of microscopy in authentication of traditional Tibetan medicinal plant Halenia elliptica

Halenia elliptica D. Don, a popularly used ethnodrug from Qinghai-Tibetan plateau, was studied to reveal the indispensable morphoanatomic details. The fixed, sectioned, and stained plant materials as well as the epidermis, powder, and maceration materials were studied using light microscope according to the usual microscopic techniques. The results of the microscopic features were systematic described and illustrated. In the root, an endodermal cell was divided into 8-16-22 and 38-50-62 daughter cells in transverse section and in face view, respectively, and 9-11-13 phloem strands were present in primary structure; in the stem, stone cells were observed in the cortex, pericycle, and external phloem while 17-19-21 internal phloem strands were present in an incontinuous ring; in the pedicel, 8-10-12 internal phloem strands were observed to form an incontinuous ring; anisocytic and anomocytic stomata were present in leaf and sepal epidermis; pollen grain was with three germinal apertures and furrows; a few tracheids, a large number of spiral vessels, and various fibers were observed. Also, semiquantitative and quantitative micrographic parameter tables were simultaneously presented. Further, the key authentication parameters were concluded. The study indicated that light microscopy and related techniques could be unambiguously applied to the authentication of Halenia elliptica.     

Morphological specializations of the epidermis of an angler catfish Chaca chaca (Siluriformes,Chacidae) in relation to its ecological niche: A scanning electron microscopic investigation

The present work was undertaken with the aim to deduce morphological adaptations in skin of an angler catfish Chaca chaca by means of scanning electron microscopy. The fish is nocturnal, bottom dwelling, sluggish, ambush predator, lives in sand, mud, or soft substrates often buried and camouflaged for protection and to feed. The surface of the epidermis is covered with polygonal epithelial cells, each having surface relief of microridges forming intricate patterns. In between epithelial cells irregularly distributed mucous cell openings, randomly distributed epidermal specialized structures, taste buds, and neuromasts are discernible. The epidermal specialized structures are keratinized. These are either irregularly the rounded elevated plaque like or the cone shaped structures. The superficial keratinized cells could frequently be discernible exfoliated at the surface. At intervals, characteristic epidermal projections could be observed. Surface of these projections at intervals is differentiated into short stumpy protuberances, each bearing a taste bud at its summit. Further, near the basal portion of these epidermal projections, conical, or rounded plaque like epidermal specialized structures are also discernible. The surface sculpture of the skin of Chaca chaca is associated with the structural and functional significance and physiological adaptations of the epidermis with respect to its ecological niche.