The development of accurate chemical kinetic models capable of predicting the combustion of methane and dimethyl ether in common combustion environments such as compression ignition engines and gas turbines is important as it provides valuable data and understanding of these fuels under conditions that are difficult and expensive to study in the real combustors. In this work, both experimental and chemical kinetic model-predicted ignition delay time data are provided covering a range of conditions relevant to gas turbine environments (T = 600 − 1600 K, p = 7 − 41 atm, φ = 0.3, 0.5, 1.0, and 2.0 in 'air' mixtures). The detailed chemical kinetic model (Mech 56.54) is capable of accurately predicting this wide range of data, and it is the first mechanism to incorporate high-level rate constant measurements and calculations where available for the reactions of DME. This mechanism is also the first to apply a pressure-dependent treatment to the low-temperature reactions of DME. It has been validated using available literature data including flow reactor, jet-stirred reactor, shock-tube ignition delay times, shock-tube speciation, flame speed, and flame speciation data. New ignition delay time measurements are presented for methane, dimethyl ether, and their mixtures; these data were obtained using three different shock tubes and a rapid compression machine. In addition to the DME/CH 4 blends, high-pressure data for pure DME and pure methane were also obtained. Where possible, the new * address:

The role of endogenous lipid in the provision of energy during in vitro maturation of immature porcine oocytes has been studied. Fluorescence resonance energy transfer (FRET) acceptor bleaching methods have been used to examine mitochondrial:lipid droplet co-localisation in live oocytes. FRET experiments demonstrate whether organelles are within the FRET-distance (i.e. 6-10 nm), thus showing true association on a molecular scale. Immature and in vitro-matured porcine oocytes were stained with Mitotracker Green (MTG; mitochondria) and Nile Red (NR; lipid droplets). The data indicated sufficient overlap between MTG emission and NR excitation to support a FRET reaction and that mitochondria and lipid droplets were sufficiently co-localised for a FRET reaction to occur. When NR-stained lipid droplets were specifically bleached, a significant increase in the MTG signal in stained mitochondria was observed (FRET efficiency, EZ22.2G3.18%). These results strongly suggest a metabolic role for lipid metabolism during oocyte maturation. This conclusion was reinforced by the use of inhibitors of fatty acid b-oxidation, methyl palmoxirate or mercaptoacetate, exposure to which during oocyte maturation led to developmental failure post-fertilisation. These data provide strong evidence that MTG and NR can act as a FRET pair and that in porcine oocytes, mitochondria and lipid droplets lie within 6-10 nm of each other, indicating association on a molecular scale. The findings also suggest that endogenous triglycerides play an important role in energy metabolism during porcine in vitro maturation. Reproduction (2006) 132 829-837

Fluorescence microscopy of GFP-tagged proteins is a fundamental tool in cell biology, but without seeing the structure of the surrounding cellular space, functional information can be lost. Here we present a protocol that preserves GFP and mCherry fluorescence in mammalian cells embedded in resin with electron contrast to reveal cellular ultrastructure. Ultrathin in-resin fluorescence (IRF) sections were imaged simultaneously for fluorescence and electron signals in an integrated light and scanning electron microscope. We show, for the first time, that GFP is stable and active in resin sections in vacuo. We applied our protocol to study the subcellular localisation of diacylglycerol (DAG), a modulator of membrane morphology and membrane dynamics in nuclear envelope assembly. We show that DAG is localised to the nuclear envelope, nucleoplasmic reticulum and curved tips of the Golgi apparatus. With these developments, we demonstrate that integrated imaging is maturing into a powerful tool for accurate molecular localisation to structure.

SummaryThe fluid mosaic model of membrane structure has been revised in recent years as it has become evident that domains of different lipid composition are present in eukaryotic and prokaryotic cells. Using membrane binding fluorescent dyes, we demonstrate the presence of lipid spirals extending along the long axis of cells of the rod-shaped bacterium Bacillus subtilis. These spiral structures are absent from cells in which the synthesis of phosphatidylglycerol is disrupted, suggesting an enrichment in anionic phospholipids. Green fluorescent protein fusions of the cell division protein MinD also form spiral structures and these were shown by fluorescence resonance energy transfer to be coincident with the lipid spirals. These data indicate a higher level of membrane lipid organization than previously observed and a primary role for lipid spirals in determining the site of cell division in bacterial cells.

Cell imaging often relies on synthetic or genetic fluorescent labels, to provide contrast which can be far from ideal for imaging cells in their in vivo state. We report on the biological application of a, label-free, high contrast microscopy technique known as ptychography, in which the image producing step is transferred from the microscope lens to a high-speed phase retrieval algorithm. We demonstrate that this technology is appropriate for label-free imaging of adherent cells and is particularly suitable for reporting cellular changes such as mitosis, apoptosis and cell differentiation. The high contrast, artefact-free, focus-free information rich images allow dividing cells to be distinguished from non-dividing cells by a greater than two-fold increase in cell contrast, and we demonstrate this technique is suitable for downstream automated cell segmentation and analysis.

In this study, we investigated whether changes in mitochondrial abundance, ultrastructure and activity are involved in the respiratory cold acclimation response in leaves of the cold-hardy plant Arabidopsis thaliana . Confocal microscopy [using plants with green fluorescence protein (GFP) targeted to the mitochondria] and transmission electron microscopy (TEM) were used to visualize changes in mitochondrial morphology, abundance and ultrastructure. Measurements of respiratory flux in isolated mitochondria and intact leaf tissue were also made. Warm-grown (WG, 25/ 20 ∞ C day/night), 3-week cold-treated (CT) and cold-developed (CD) leaves were sampled. Although CT leaves exhibited some evidence of acclimation (as evidenced by higher rates of respiration at moderate measurement temperatures), it was only the CD leaves that were able to reestablish respiratory flux within the cold. Associated with the recovery of respiratory flux in the CD leaves were: (1) an increase in the total volume of mitochondria per unit volume of tissue in epidermal cells; (2) an increase in the ratio of cristae to matrix within mesophyll cell mitochondria; and (3) an increase in the capacity of the energyproducing cytochrome pathway in mitochondria isolated from whole leaf homogenates. Regardless of growth temperature, we found that contrasting cell types exhibited distinct differences in mitochondrial ultrastructure, morphology and abundance. Collectively, our data demonstrated the diversity and tissue-specific nature of mitochondrial responses that underpin respiratory acclimation to the cold, and revealed the heterogeneity of mitochondrial structure and abundance that exists within leaves.

An understanding of the ignition and oxidation characteristics of propanol, as well as other alcohols, is important toward the development and design of combustion engines that can effectively utilize bioderived and bioblended fuels. Building upon a database for “first-generation” alcohols including methanol and ethanol, the ignition characteristics of the two isomers of propanol (n-propanol and iso-propanol) have been studied in a shock tube. Ignition delay times for propanol/oxygen/argon mixtures have been measured behind reflected shock waves at temperatures ranging from approximately 1350 to 2000 K and a pressure of 1 atm. Equivalence ratios of 0.5, 1.0, and 2.0 have been used. Pressure measurements and CH* emissions were used to determine ignition delay times. The influences of equivalence ratio, temperature, and mixture strength on ignition delay have been characterized and compared to the behavior seen with a newly developed detailed kinetic mechanism. The overall trends are captured fairly well by the mechanism, which include a greater level of reactivity for the n-propanol mixtures relative to iso-propanol at the conditions used in this study.

Label-free imaging uses inherent contrast mechanisms within cells to create image contrast without introducing dyes/labels, which may confound results. Quantitative phase imaging is label-free and offers higher content and contrast compared to traditional techniques. High-contrast images facilitate generation of individual cell metrics via more robust segmentation and tracking, enabling formation of a label-free dynamic phenotype describing cell-to-cell heterogeneity and temporal changes. Compared to population-level averages, individual cell-level dynamic phenotypes have greater power to differentiate between cellular responses to treatments, which has clinical relevance e.g. in the treatment of cancer. Furthermore, as the data is obtained label-free, the same cells can be used for further assays or expansion, of potential benefit for the fields of regenerative and personalised medicine.