Purification and characterization of a nuclear DNA-binding phosphoprotein in fetal and tumor tissues

The basic nonhistone phosphoprotein 110/8.4 (M.W. X 10(-3)/pI) was found in 0.35 M NaCl nuclear extracts of four tumor tissues, i.e., fast-growing Novikoff hepatoma, Morris hepatoma 3924A, HeLa cells, and Namalwa cells; it was also found in fetal rat liver. This protein was not detected in normal or regenerating liver and thus may represent an "oncofetal" protein of potential interest as a cancer "marker." Protein 110/8.4 was purified approximately 4000- to 5000-fold under nondenaturing condition from 0.35 M NaCl nuclear extracts of Novikoff hepatoma cells or Namalwa cells by ammonium sulfate fractionation, calcium phosphate gel treatment, and phosphocellulose chromatography. Sodium dodecyl sulfate:polyacrylamide gel electrophoresis of the purified native protein revealed a single polypeptide chain with a molecular weight of approximately 110,000. The pI of the protein was estimated to be 8.4 by nonequilibrium pH gradient electrophoresis in 9 M urea; accordingly, this protein was designated 110/8.4. Amino acid analysis showed that Protein 110/8.4 had an acidic:basic amino acid ratio of 1.25 and a high lysine and serine content; approximately 20% of the serine residues were found to be phosphorylated. Hydrazinolysis indicated that the carboxyl-terminal amino acid was serine; the amino terminus appeared to be blocked. Binding of Protein 110/8.4 to DNA was studied by the nitrocellulose filter assay. High-affinity binding occurred at ionic strength equal to or below 0.15 M.     

Detection of Borrelia burgdorferi-specific CD8+ cytotoxic T cells in patients with Lyme arthritis

T cells are believed to play an important role in the pathogenesis of Lyme arthritis (LA), an inflammatory joint disease caused by the spirochete Borrelia burgdorferi (Bb). The presence or absence of certain Bb-specific CD4+ T helper cells has been associated with prognosis. Since recent observations suggested the activation of CD8+ T cells during infection with Bb, we searched for CD8+ cytotoxic T lymphocytes in patients with LA. CD8+ T cell lines were generated from peripheral blood and synovial fluid of five patients with LA. In addition, CD8+ T cells were expanded by Ag-specific stimulation in bulk cultures. A cytotoxicity assay was established using target cells infected with recombinant vaccinia viruses expressing the borrelial proteins outer surface protein (Osp) A, OspB, or flagellin. We found Bb-specific CTL lines derived from the peripheral blood of three patients with LA with specificity for flagellin, OspA, and OspB. All Bb-specific CTL lines were CD3+, CD8+, and TCRalphabeta, and cytotoxic activity was HLA class I restricted. Moreover, CD8+ T cells expanded by Ag-specific stimulation in vitro demonstrated Bb-specific and HLA class I-restricted lysis toward individual borrelial proteins. Interestingly, Bb-specific lytic activity was only detected in patient samples obtained after the disappearance of arthritis. We report the detection of Bb-specific cytotoxic CD8+ T cells in patients with LA. The induction of specific CD8+ T cells may play an important role in disease control and may have important bearings for the development of effective vaccines against Lyme borreliosis.     

Grain-boundary penetration of type 316 stainless steel exposed to cesium or cesium and tellurium

When 20 pct cold-worked Type 316 stainless steel is exposed to Cs at 700°C under controlled oxygen-chemical potential environment, Cs penetration into the stainless steel grain boundaries occurs at oxygen potentials ΔG_o2 ≥ -96 kcal per mole. At lower oxygen potentials (~ΔG_o2 ≤ —110 kcal per mole), no corrosion occurs. Under the same experimental conditions, when the stainless steel is exposed to Cs:Te (2:1, atomic), corrosion occurs and penetration morphology appears to depend strongly on the oxygen-potential environment. The stainless steel suffers intergranular corrosion by Te (in the presence of Cs-Te) under conditions where chromium oxidation is not expected to occur. The kinetics of grain-boundary penetration by Te have been studied at temperatures between 550 and 700°C. The depth of the penetrated zone varies as (time)^1/2, and the process has an activation energy of 34 kcal per mole. The results are discussed, and the effects of stainless steel microstructure and externally applied stress on corrosion reactions are also described.     

Synthesis of active carbon-based catalysts by chemical vapor infiltration for nitrogen oxide conversion

Direct reduction of nitrogen oxides is still a challenge. Strong efforts have been made in developing noble and transition metal catalysts on microporous support materials such as active carbons or zeolites. However, the required activation energy and low conversion rates still limit its breakthrough. Furthermore, infiltration of such microporous matrix materials is commonly performed by wet chemistry routes. Deep infiltration and homogeneous precursor distribution are often challenging due to precursor viscosity or electrostatic shielding and may be inhibited by pore clogging. Gas phase infiltration, as an alternative, can resolve viscosity issues and may contribute to homogeneous infiltration of precursors. In the present work new catalysts based on active carbon substrates were synthesized via chemical vapor infiltration. Iron oxide nano clusters were deposited in the microporous matrix material. Detailed investigation of produced catalysts included nitrogen oxide adsorption, X-ray diffraction, scanning electron microscopy and energy-dispersive X-ray spectroscopy. Catalytic activity was studied in a recycle flow reactor by time-resolved mass spectrometry at a temperature of 423 K. The infiltrated active carbons showed very homogeneous deposition of iron oxide nano clusters in the range of below 12 to 19 nm, depending on the amount of infiltrated precursor. The specific surface area was not excessively reduced, nor was the pore size distribution changed compared to the original substrate. Catalytic nitrogen oxides conversion was detected at temperatures as low as 423 K.     

Nachweis und Charakterisierung von Clostridium perfringens mittels real-time-PCR

Clostridium perfringens is a Gram-positive, anaerobic bacterium. The strains are classified into five types (type A to E), depending on the ability to produce alpha-, beta-, epsiloon and iota-toxin, but food-poisonings are mostly caused by C. perfringens type A isolates. Ingestion of contaminated food is followed by gastrointestinal disease, when enzyme-resistant C. perfringens enterotoxins (CpE) are set free during sporulation. In most cases the bacterium has to grow up to more than 10⁶ cfu/g food to cause a gastrointestinal disease (BVET, 2005; BfR, 2005). Cultural methods, normally used for the detection of C. perfringens, are not able to differentiate isolates into the five different toxin types. This is the reason, why the detection of C. perfringens in food under the present legal regulations can only used as an indication for inadequate production hygiene or a toxin-infection with contaminated food as source. For the detection of the toxin genes to differentiate C. perfringens in five different toxin types, three duplex real-time-PCR assays for the routine diagnostic were developed and validated. The assays can be used for quick detection and easy classification of C. perfringens isolates and as a rapid screening-system in suspected cases of C. perfringens food-poisonings.     

Quantification of lupine (Lupinus angustifolius) in wheat flour using real-time PCR and an internal standard material

In the European Union, labelling of the 14 food allergens listed in Annex IIIa of Directive 2000/13/EC is mandatory. The implementation of upper limits for these allergens is under discussion. Therefore, quantitative analytical methods will be needed to verify compliance with regulatory requirements and to provide an improved basis for the legal assessment of allergen labelling. In this study, the lupine flour content in wheat flours was determined using real-time PCR and statice seeds as internal standard material. The method proved to be applicable to the quantification of lupine contents from 1 to 10?mg/kg, which is in the range relevant for allergic consumers.     

European inter-laboratory comparison of high pressure CO2 sorption isotherms. I: Activated carbon

In order to assess and improve the quality of high-pressure sorption isotherms of carbon dioxide (CO₂) on coals, an inter-laboratory study (“Round Robin”) has been conducted among four European research laboratories. In a first round of measurements, excess sorption isotherms were determined on Filtrasorb 400 (F400) activated carbon at 318 K using the manometric (TU Delft and RWTH Aachen University) and the gravimetric (FP Mons and INERIS) method up to 16 MPa. The study shows that CO₂ sorption in the supercritical range can be determined accurately with both gravimetric and manometric equipment but requires thorough optimization of instrumentation and measuring as well as proper sample preparation procedures. For the characterization of the activated carbon F400, which we used as benchmark, we have determined a surface area of 1063 m² g⁻¹, and Dubinin-Radushkevich (DR) micropore volume of 0.51 cm³ g⁻¹. Additionally, we analysed the elementary near-surface composition by energy dispersive X-ray spectroscopy (EDX). To characterise the bulk composition of the F400 activated carbon, a proximate and ultimate analysis was performed.The observed excess sorption maxima around 5 MPa have values around 8.0 mol kg⁻¹, which are consistently higher (by upto 0.8 mol kg⁻¹) than literature data.