A Rapid and Efficient Immunoenzymatic Assay to Detect Receptor Protein Interactions: G Protein-Coupled Receptors

G protein-coupled receptors (GPCRs) represent one of the largest families of cell surface receptors, and are the target of at least one-third of the current therapeutic drugs on the market. Along their life cycle, GPCRs are accompanied by a range of specialized GPCR-interacting proteins (GIPs), which take part in receptor proper folding, targeting to the appropriate subcellular compartments and in receptor signaling tasks, and also in receptor regulation processes, such as desensitization and internalization. The direction of protein-protein interactions and multi-protein complexes formation is crucial in understanding protein function and their implication in pathological events. Although several methods have been already developed to assay protein complexes, some of them are quite laborious, expensive, and, more important, they do not generate fully quantitative results. Herein, we show a rapid immunoenzymatic assay to quantify GPCR interactionswith its signaling proteins. The recently de-orphanized GPCR, GPR17, was chosen as a GPCR prototype to optimize the assay. In a GPR17 transfected cell line and primary oligodendrocyte precursor cells, GPR17 interaction with proteins involved in the typical GPCR regulation, such as desensitization and internalization machinery, was investigated. The obtained results were validated by co-immunoprecipitation experiments, confirming this new method as a rapid and quantitative assay to study protein-protein interactions.     

Evaluation of the false-positive enteroviral plaque phenomenon occurring in sewage samples

Several laboratories monitoring for enteroviruses in wastewater have reported a high percentage of false-positive viral plaques. This article discusses the issue of possible false evaluation of viral plaque determinations in wastewater, and reviews the preventive measures taken in several wastewater studies conducted in our laboratory, which minimized this phenomenon. Our results showed that with the procedures described, no false-positive plaques were found.     

A novel tunable-complexity turbo-soft detector for high-throughputHDSL applications over ISI channels with crosstalk

We present a novel iterative (turbo) receiver with tunable complexity for reliable detection of (uncoded) payload data transmitted over long intersymbol interference (ISI) channels affected by crosstalk, as those typically encountered in emerging HDSL2 services standardized by ANSI T1.418 recommendation. The proposed receiver combines in an original way "minimum mean square error (MMSE) estimation principle," "turbo-processing principle" and "crosstalk-prediction principle" for achieving both suboptimal maximum a posteriori probability channel equalization and reliable soft-mitigation of ISI tail plus crosstalk. More in detail, according to the turbo-processing principle, at each iteration suitable extrinsic information is extracted from the equalizing and interference-canceling modules and used as "a priori information" for the next iteration. Several simulation results on typical HDSL-like test-loops confirm the superiority of the proposed turbo-detector (TD) over current solutions based on conventional MMSE decision-feedback equalizers and precoders (such as the Tomlinson-Harashima precoder). The numerical tests also point out that performance of the presented TD on typical high bit-rate digital subscriber lines (HDSL) is not limited by error-floor phenomena, even at error-probabilities below 10^-7     

New chiral inhibitors of induced platelet aggregation: the enantiomeric specificity of (R)- and (S)-methyl and ethyl esters of 1-(4-[(1-hydroxycarbonyl)-ethoxy]-benzyl)-1H-1,2,3-triazole as a tool for determining their biological target

The synthesis of title enantiomers was accomplished and their biological behaviour as inhibitors of rabbit platelet aggregation process induced by ADP and arachidonic acid was determined. Structure-activity comparison with that of SM-12502 [(2R,5S)-(+) 3,5-dimethyl-2-(3-pyridyl)-thiazolidin-4-one hydrochloride] and Dazoxiben [4-[2-(1H-imidazol-1-yl)-ethoxy]-benzoic acid] allowed us to formulate the possible capability for the synthesized compounds to interact with the biological targets of the model molecules.     

Distribution of a simple shared dataspace architecture

We study a simple software architecture, in which components are coordinated by writing into and reading from a global set. This simple architecture is inspired by the industrial software architecture Splice. We present two results. First, a distributed implementation of the architecture is given and proved correct formally. In the implementation, local sets are maintained and data items are exchanged between these local sets. Next we show that the architecture is sufficiently expressive in principle. In particular, every global specification of a system's behaviour can be divided into components, which coordinate by read and write primitives on a global set only. We heavily rely on recent concepts and proof methods from process algebra.     

Enzymatic fatty ester synthesis

Fatty ester synthesis with immobilized 1,3-specific lipase fromMucor miehei is described. 1,2-Isopropylidene glycerol produced by condensation of glycerol with acetone was esterified with oleic acid in the presence of aMucor miehei lipase (Lipozyme™) to obtain 1,2-isopropylidene-3-oleoyl glycerol. The effects of various process parameters (temperature and pressure) and various ratios (enzyme/substrate) have been investigated to determine optimal conditions for the esterification process. The highest conversion of oleic acid (80% w/w) was obtained at 55°C and 0.057 bar, while the optimal addition of lipase to substrate was determined to be 0.096 g per gram of reaction mixture. The esterification can be modeled successfully as a reverse second-order reaction. Thermodynamic properties of the reaction system at 55°C and 0.057 bar also were determined. Activation energy was 20.82 kJ/mole, entropy of activation −0,26 kJ/(K mole) and free energy of activation was 103.32 kJ/mole.     

Polymerization Kinetics and Characterization of Dual Cured Polyurethane‐Acrylate Nanocomposites for Laminates

Summary: Four different types of montmorillonites have been dispersed by sonication at 50 °C into a propoxylated aromatic epoxy diacrylate oligomer to achieve interlayered or exfoliated nanocomposites. A thermally‐induced crosslinking reaction, forming a polyurethane network in the presence of 7 wt.‐% of a montmorillonite, has been promoted by addition of an allophanate modified polyisocyanurate based on hexamethylene diisocyanate. The kinetic behavior of the network formation has been studied at 25, 40 and 60 °C by following the disappearance of the isocyanate vibrational band found at 2 270 cm^?1. A tight crosslinked polyurethane acrylate network has been achieved by a subsequent dual UV curing promoted by a photoinitiator mixture (0.6 wt.‐%) added to the reactive mixture because of further reactions occur to the acrylate double bonds. The photopolymerization kinetic has been investigated on the different thermally treated polyurethane nanocomposite networks by Real Time FTIR spectroscopy monitoring the changes of the IR band at 810 cm^?1 assigned to the acrylate double bond vibrations. The influence of the different montmorillonite clays on the final nanocomposite morphology has been investigated by using XRD and SEM. Finally, the use of these mixtures as internal layer between two modified surface PET films has been also studied for the laminate production. The based‐PET laminate films have been characterized by determining the bending resistance and optical properties as a function of different nanofillers.

Bending resistance of the dual cured nanocomposite laminates containing 7 wt.‐% as a function of nanofiller types.     

Estimation of Evapotranspiration of Different-Sized Navel-Orange Tree Orchards Using Energy Balance

Crop evapotranspiration (ETc) and crop coefficient (Kco) values of four clean-cultivated navel-orange orchards that were irrigated with microsprinklers, having different canopy features (e.g., age, height, and canopy cover) were evaluated. Half-hourly values of latent heat flux density were estimated as the residual of the energy balance equation using measured net radiation (Rn), soil heat flux density (G), and sensible heat flux density (H) estimated using the surface renewal method. Hourly means of latent heat flux density (LE) were calculated and were divided by the latent heat of vaporization (L) to obtain ETc. Crop coefficients were determined by calculating the ratio Kco = ETc/ETo, with reference evapotranspiration (ETo) determined using the hourly Penman–Monteith equation for short canopies. The estimated Kco values ranged from 0.45 to 0.93 for canopy covers having between 3.5 and 70% ground shading. The Kco values were compared with Kc values from FAO 24 (reported by Doorenbos and Pruitt in 1975) and FAO 56 (reported by Allen et al. in 1998) and with Kc values from research papers that estimated reference ET from pan evaporation data using the FAO 24 method. The observed Kco values were slightly higher than Kc values for clean-cultivated orchards with high-frequency drip irrigation in Arizona and were slightly lower than for nontilled orchards in Florida. The Kco values were considerably higher than Kc values from FAO 24 and FAO 56 and were higher than Kc values from border-irrigated orchards near Valencia, Spain.