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The reliable production of large amounts of stable, high-quality proteins is a major challenge facing pharmaceutical protein biochemists, necessary for fulfilling demands from structural biology, for high-throughput screening, and for [...]

An integrated 10-pump eight-channel LC/MS system has been developed for automated high-throughput analysis of intact proteins in recombinant protein purification processes. The key features of the system include (1) a compact 10-pump HPLC module that uses two pumps to generate a binary gradient and 8 pumps to deliver the mixed gradient to eight independent flow channels; (2) a TOF mass spectrometer with an eight-channel multiplexed ESI interface, which records separate data for all eight channels over each HPLC run cycle; and (3) highly automated data processing software that allows unattended calculation of protein molecular weight (in Da) from original mass spectral data (in m/z). This system was used in the routine screening of fractions from preparative scale chromatography to monitor the purification process with the required mass accuracy and throughput. As an example, the production and purification of an acylated protein with a molecular weight of 9 kDa is described. Using this off-line approach, it is practical to fully characterize protein-containing fractions from column chromatography with an overall analytical throughput of 1 min/protein sample with minimum operator involvement.

The experiment concerning the ECLISSE method (ECR ion source coupled to a laser ion source for charge state enhancement) has been carried out by coupling a laser ion source (LIS) to the superconducting electron cyclotron resonance source (SERSE) electron cyclotron resonance (ECR) ion source with the goal to obtain intense beams of highly charged ions (cw or pulsed mode) from metal samples, especially from refractory elements. The coupling efficiency of the ion beam produced by the LIS with the ECR plasma was remarkable and the measured beam intensities were quite high. The maximum charge states, obtained with a good reproducibility, were 38+ for Ta and 41+ for Au. The highest current was obtained for 25+ and 28+ for Ta and Au ions, respectively, and it was in both cases of the order of some tens of microampere, i.e., higher than the current obtained from SERSE with other methods (i.e., evaporation and sputtering). The ion beam stability and reproducibility were both acceptable. The possibility to get a further enhancement of the available charge states was limited by charge exchange processes connected with the target ablation. The main features of the LNS facility based on a Nd:YAG (yag, yttrium aluminum garnet) laser (0.9 J/9 ns, laser power densities <5×1010 W/cm2) and on the SERSE superconducting ECRIS working in single frequency mode (18 GHz) will be shown in the following. The analysis of the results and the way to get rid of the existing limits will be also presented and discussed. [ABSTRACT FROM AUTHOR]

Ansamycins reduced to their dihydroquinones in the presence of common reducing agents in vitro have approximately 40-fold greater affinity than the corresponding oxidized quinones. The in vivo accumulation of the high-affinity dihydroquinone ansamycins in tumor cells contributes to the antitumor activity of these compounds and alters the understanding of the active species driving the efficacy of this class of compounds.

A pulsed Nd:Yag laser, at intensities of the order of 1010 W/cm2, is employed to irradiate different thick metallic targets (Ti, Fe, Ag, and Ni) placed in vacuum. The obtained non-equilibrium plasmas are investigated with various analytical techniques. An electrostatic ion energy analyzer and different ion collectors are employed to monitor in situ the ions ejected from the plasma and to determine the core plasma temperature, the ion energy distributions and the ion angular emission. An optical spectrometer is employed to analyze the plasma corona emitted light vs. wavelength and to identify the emitted characteristic lines. The optical spectroscopy permitted to evaluate the electron temperatures and densities. Results show strong temperature and density gradients occurring in the laser-generated plasma plume. [ABSTRACT FROM AUTHOR]

Physical analyses by laser ablation coupled to mass quadrupole spectrometry (LAMQS), energy dispersive X-ray fluorescence (EDX) induced by electron beam, scanning electron microscopy (SEM) and surface profilometry analysis (SPA) are applied to the investigation of two silver tetradrachms from Messana, in order to compare their elemental composition and structure. Quantitative analysis of the elemental composition and of the silver isotopic ratios have been carried out analyzing the surface patina of the two samples. Significant differences in the sulfur, chlorine and copper content, in the isotopic ratios and in the morphological aspects have been measured. The obtained results are presented and discussed from the point of view of the physical techniques useful to establish the differences between apparently true and false coins. [ABSTRACT FROM AUTHOR]

A Nd:YAG laser operating in second harmonic (532 nm), 3 ns pulse duration, 150 mJ pulse energy, and 10 Hz repetition rate, is employed to irradiate Al2O3 target placed in high vacuum. The produced plasma is investigated by an ion collector used in time-of-flight configuration and by a mass quadrupole spectrometer, in order to determine the equivalent plasma temperature and the atomic and molecular composition. Pulsed laser deposition technique has been used to produce thin films on different substrates placed close to the target. Different surface analyses, such as energy dispersive X-ray fluorescence (EDXRF), X-ray photoelectron spectroscopy (XPS) and surface profilometry are employed to characterize the produced films. Measurements of ablation yield, plasma equivalent temperature, acceleration voltage and characterization of grown thin films are presented and discussed. [ABSTRACT FROM AUTHOR]

A Nd–YAG laser operating at 532 nm with a maximum intensity of 10 10 W/cm 2 was used to ablate aluminium and tantalum targets placed in vacuum. A mass quadrupole spectrometer (MQS) at high sensitivity, operating in the range of 1–300 amu, with a resolution better than 1 amu, was used to analyse the atomic emission produced by the laser ablation. The neutrals' emission produced by laser-generated plasma at INFN-LNS was investigated in terms of temperature, ablation yield, angular distribution and velocity. The neutrals' detection through the MQS permitted to measure the mass energy distribution. Results demonstrate that the maximum temperatures of the neutral species are of the order of 100 eV. The angular emission of neutrals is peaked along the normal to the target surface, as it was observed for the ions; the ablation yield increases suddenly at low laser intensity and decreases at high laser intensity, owing to the higher ionization processes; the flow velocity follows the adiabatic expansion of the plasma in vacuum and it is of the order of 10 4 m/s. Measurements will be presented and discussed, according to the available models. [ABSTRACT FROM AUTHOR]