Extraction methods of red blood cell membrane proteins for Multidimensional Protein Identification Technology (MudPIT) analysis

Since red blood cells (RBCs) lack nuclei and organelles, cell membrane is their main load-bearing component and, according to a dynamic interaction with the cytoskeleton compartment, plays a pivotal role in their functioning. Even if erythrocyte membranes are available in large quantities, the low abundance and the hydrophobic nature of cell membrane proteins complicate their purification and detection by conventional 2D gel-based proteomic approaches. So, in order to increase the efficiency of RBC membrane proteome identification, here we took advantage of a simple and reproducible membrane sub-fractionation method coupled to Multidimensional Protein Identification Technology (MudPIT). In addition, the adoption of a stringent RBC filtration strategy from the whole blood, permitted to remove exhaustively contaminants, such as platelets and white blood cells, and to identify a total of 275 proteins in the three RBC membrane fractions collected and analysed. Finally, by means of software for the elaboration of the great quantity of data obtained and programs for statistical analysis and protein classification, it was possible to determine the validity of the entire system workflow and to assign the proper sub-cellular localization and function for the greatest number of the identified proteins.     

Investigation of the interface in silica-encapsulated liposomes by combining solid state NMR and first principles calculations

In the context of nanomedicine, liposils (liposomes and silica) have a strong potential for drug storage and release schemes: such materials combine the intrinsic properties of liposome (encapsulation) and silica (increased rigidity, protective coating, pH degradability). In this work, an original approach combining solid state NMR, molecular dynamics, first principles geometry optimization, and NMR parameters calculation allows the building of a precise representation of the organic/inorganic interface in liposils. {(1)H-(29)Si}(1)H and {(1)H-(31)P}(1)H Double Cross-Polarization (CP) MAS NMR experiments were implemented in order to explore the proton chemical environments around the silica and the phospholipids, respectively. Using VASP (Vienna Ab Initio Simulation Package), DFT calculations including molecular dynamics, and geometry optimization lead to the determination of energetically favorable configurations of a DPPC (dipalmitoylphosphatidylcholine) headgroup adsorbed onto a hydroxylated silica surface that corresponds to a realistic model of an amorphous silica slab. These data combined with first principles NMR parameters calculations by GIPAW (Gauge Included Projected Augmented Wave) show that the phosphate moieties are not directly interacting with silanols. The stabilization of the interface is achieved through the presence of water molecules located in-between the head groups of the phospholipids and the silica surface forming an interfacial H-bonded water layer. A detailed study of the (31)P chemical shift anisotropy (CSA) parameters allows us to interpret the local dynamics of DPPC in liposils. Finally, the VASP/solid state NMR/GIPAW combined approach can be extended to a large variety of organic-inorganic hybrid interfaces.     

Development and characterisation of an XeCl excimer laser-generated soft-X-ray plasma source and its applications

Summary A large-aperture, long-pulse XeCl excimer laser has been used to generate a soft-X-ray plasma source. Several laser optical configurations have been employed to optimise X-ray emission, including positive-branch unstable resonators and injection by seeding the gain region of the laser with a small commercial excimer laser, resulting in power densities in the range 10¹²–10¹⁴ W cm⁻². The characteristics of the plasma source for each different laser configuration and for different target materials are investigated. The most suitable source conditions (spectral energy distribution, time duration, etc.) for specific applications are discussed. Supported by the Royal Society, UK. Supported by INFM, Italy.     

Flame atomic absorption analysis of gold in jewelry samples

Summary A new procedure is proposed for the determination of gold by flame atomic absorption in jewelry samples. The method is based on the acid digestion of 10 mg of sample in a microwave oven and on the on-line dilution, using a three-way valve. The dynamic range of the technique is extended to 100 mg/l gold solutions and using a weighted bracketing method for the treatment of the results, accuracy errors lower than 2% can be found in the analysis of real samples as compared with those obtained by fire assay. The method is very fast and implies only little damage to the jewelry samples.     

Study of a positive corona discharge in argon at different pressures

The corona discharge in argon at atmospheric pressure has been studied by means of a 2D model. The reduced characteristic derived from the experimental data has been described by linear regressions for the different pressures and the two studied inter-electrode distances thus confirming the validity of Townsend's approximation also in case of point to plane configuration and argon as process gas. The model validated this hypothesis which has been attributed to the minor influence of space charge in the ionization zone. Its effect is, on the other hand, more significant in the drift zone where the electric field is greatly enhanced, leading, for higher currents, to the formation of a spark gap. Electron and ion distributions allow the influence of structural (electrode configurations and distance) and operative (pressure and discharge current) parameters to be evaluated including the current loss due to diffusion through different confining boundaries.     

Conformational analysis XX—13C NMR studies of saturated heterocycles 5—substituent effects on the 13C chemical shifts of methyl substituted 1,3-dithiolanes

The ¹³C NMR chemical shifts for 1,3-dithiolane and 13 methyl substituted derivatives are reported. Substituent effects are derived and compared with those for cyclopentanes and 1,3-dioxolanes. The magnitude and variety of the substituent effects are best explained with the aid of a half-chair conformation where the S-1? C-2? S-3 plane passes between C-4 and C-5.