Capillary Electrophoretic Analysis Reveals Subcellular Binding between Individual Mitochondria and Cytoskeleton

Interactions between the cytoskeleton and mitochondria are essential for normal cellular function. An assessment of such interactions is commonly based on bulk analysis of mitochondrial and cytoskeletal markers present in a given sample, which assumes complete binding between these two organelle types. Such measurements are biased because they rarely account for nonbound "free" subcellular species. Here we report on the use of capillary electrophoresis with dual laser induced fluorescence detection (CE-LIF) to identify, classify, count, and quantify properties of individual binding events of the mitochondria and cytoskeleton. Mitochondria were fluorescently labeled with DsRed2 while F-actin, a major cytoskeletal component, was fluorescently labeled with Alexa488-phalloidin. In a typical subcellular fraction of L6 myoblasts, 79% of mitochondrial events did not have detectable levels of F-actin, while the rest had on average ～2 zmol of F-actin, which theoretically represents a ～2.5 μm long network of actin filaments per event. Trypsin treatment of L6 subcellular fractions prior to analysis decreased the fraction of mitochondrial events with detectable levels of F-actin, which is expected from digestion of cytoskeletal proteins on the surface of mitochondria. The electrophoretic mobility distributions of the individual events were also used to further distinguish between cytoskeleton-bound from cytoskeleton-free mitochondrial events. The CE-LIF approach described here could be further developed to explore cytoskeleton interactions with other subcellular structures, the effects of cytoskeleton destabilizing drugs, and the progression of viral infections.     

Calcium sulphide formation in fluidized bed boilers

Analyses of samples of bed ash from a stationary fluidized bed boiler show the presence of calcium sulphide. In some samples, half of the total sulphur was present as sulphide. The samples containing CaS were obtained under unstaged conditions and with a high excess air ratio, 1.3 to 1.4. The samples were taken after a stop in the limestone addition, i.e. at high SO₂ emissions of about 1000 mL/m³ (ppm). No CaS was found during limestone addition when the SO₂ emission was 300–400 mL/m³. This indicates that formation of large amounts of CaS may be initiated as the SO₂ concentration exceeds some critical level.     

Effect of thiourea on thiuram-accelerated sulfur vulcanization and its significance for vulcanization mechanism

Tetramethylthiuram disulfide-accelerated sulfur vulcanization of natural rubber has been investigated at temperatures from 100°C to 140°C. Continuous measurements in a Vuremo curemeter were used to estimate the extent of crosslinking, which was plotted against cure time. The linearized forms of the cure curves clearly show that at lower cure temperatures, the course of vulcanization differs significantly from the first-order rate law. These digressions were removed by the addition of thiourea, which simultaneously speeds up the course of vulcanization and diminishes its activation energy on the one hand but, on the other hand, diminishes the ultimate extent of crosslinking. On the basis of the above results, the mechanism of thiuram-accelerated sulfur vulcanization, in which ionic and radical reactions take place, is discussed.     

Dielectric response and structure of amorphous hydrogenated carbon films with nitrogen admixture

The optical properties and structure of a-C:H films were modified by addition of nitrogen into the CH₄/H₂ deposition mixture. Three films prepared in capacitively coupled rf discharge were compared: (a) hydrogenated diamond like carbon film with hydrogen content of 34% and indentation hardness of 21.7 GPa, (b) hard a-C:H:N film with nitrogen content of 13% and indentation hardness of 18.5 GPa and (c) soft a-C:H:N film with nitrogen content of 10% and indentation hardness of 6.7 GPa. It is shown how the parametrized density of states model describing dielectric response of electronic interband transitions can be applied to modified a-C:H:N and how it can be combined with correct treatment of transmittance measured in infrared range using additional Gaussian peaks in joint density of phonon states. This analysis resulted in determination of film dielectric function in wide spectral range (0.045-30 eV) and provided also information about the density of states of valence and conduction bands and lattice vibrations.     

Contribution to the mechanism of sulphur-free thiuram vulcanisation

The influence of the concentration of tetramethylthiuram disulphide on the kinetics and degree of the sulphur-free vulcanisation of pale crepe in the absence and in the presence of thiourea has been investigated. Rate and degree of the vulcanisation increase with concentration of tetramethylthiuram disulfide to a certain limit. Exceeding this limit the values of the rate constant and the crosslinking density decrease. Due to the presence of thiourea the rate of the vulcanisation is increased and the final degree of vulcanisation is decreased. This effect of thiourea also depends on the concentration of tetramethylthiuram disulphide. On the basis of the above results the mechanism of the tetramethylthiuram disulphide vulcanisation is discussed.     

Theoretical Structure Determination of a Complex Material: κ-Al2O3

The atomic and electronic structures of kappa-Al₂O₃ are determined using theoretical first-principles techniques based on density-functional theory (DFT), plane waves, and pseudopotentials. The obtained structure is confirmed by analysis of powder X-ray diffraction data. The structure is orthorhombic with oxygen ions in close-packed ABAC stacking and aluminum ions occupying both tetrahedral (1/4) and octahedral (3/4) interstitial sites. A growth model for chemical vapor deposition of kappa-Al₂O₃ is proposed based on the atomic structure. Calculated electronic structure and charge density yield a band gap of 5.3 eV and a high ionic character of the bonds. The study shows the applicability of DFT-based methods to complex and metastable materials.     

Stabilization/solidification of galvanic sludges by asphalt emulsions

A combination of two-aqueous asphalt emulsions was proposed for stabilization/solidification treatment of galvanic sludges prior to landfilling. The presented procedure comprises mixing the galvanic sludge with a slow setting nonionic asphalt emulsion and subsequently forming a secondary asphalt barrier by means of a rapid setting anionic asphalt emulsion. The method was tested on four samples of galvanic sludge from various galvanizing plants, with various water and pollutant contents. Stabilization efficiency was assessed by water-leaching test, TCLP test and by determining ecotoxicity of leachates. Leachate parameters exhibited very low values and favorable results of ecotoxicological tests indicate high efficiency of the developed procedure for galvanic sludge disposal.     

Stabilization/solidification of munition destruction waste by asphalt emulsion

Destruction of discarded military munitions in an explosion chamber produces two fractions of hazardous solid waste. The first one is scrap waste that remains in the chamber after explosion; the second one is fine dust waste, which is trapped on filters of gas products that are exhausted from the chamber after explosion. The technique of stabilization/solidification of the scrap waste by asphalt emulsion is described in this paper. The technique consists of simple mixing of the waste with anionic asphalt emulsion, or two-step mixing of the waste with cationic asphalt emulsion. These techniques are easy to use and the stabilized scrap waste proves low leachability of contained heavy metals assessed by TCLP test. Hence, it is possible to landfill the scrap waste stabilized by asphalt emulsion. If the dust waste, which has large specific surface, is stabilized by asphalt emulsion, it is not fully encapsulated; the results of the leaching tests do not meet the regulatory levels. However, the dust waste solidified by asphalt emulsion can be deposited into an asphalted disposal site of the landfill. The asphalt walls of the disposal site represent an efficient secondary barrier against pollutant release.