Absolute quantification of proteins in solutions and in polyacrylamide gels by mass spectrometry

A combination of nanoelectrospray tandem mass spectrometry and (18)O-labeled peptide internal standards was applied for the absolute quantification of proteins from their in-solution and in-gel tryptic digests. Although absolute quantification from in-solution digests was accurate, we observed that in-gel digestion compromised the quantification accuracy by affecting the recovery of individual peptides and, therefore, the provided estimates might be strongly influenced by the selection of reference peptides. Under optimized experimental conditions, it was possible to provide a semiquantitative estimate of the absolute amount of gel separated proteins within better than 50% error margin.     

Cooperative Coding for OFDM Systems

In this paper, we study Orthogonal Frequency Division Multiplexing (OFDM) systems with cooperative coding over frequency selective Rayleigh fading channels. We derive the pairwise error probability (PEP) for the block-fading OFDM channel model. We use the derived PEP to get an upper bound on the frame error probability for the coded cooperative OFDM system. This bound is then utilized in the study of the diversity and coding gains achievable through cooperative coding in OFDM systems for various inter-user channel qualities. We consider the design of cooperative convolutional codes based on the principle of overlays and provide simulation results for different cooperation scenarios. We observe significant gains over conventional non-cooperative OFDM systems. Finally, based on some simple approximations, we provide guidelines for the choice of partners in coded cooperative OFDM systems.     

A discrete‐components millimeter‐wave satellite beacon receiver for Q‐band propagation experiment

Ever increasing bandwidth requirements in satellite communications continuously push the frequency limits. Q‐ band (33 – 50 GHz) is the next frequency band to be populated; however, at these frequencies, the Earth's troposphere (weather) profoundly alters the radio propagation conditions. Therefore, in order to properly plan radio links, accurate statistical models of radio channels are necessary. These statistical models are built upon empirical data, ie, measurements, which are furthermore used to design appropriate Propagation Impairment Mitigation Techniques. The Q‐ band lacks such data, hence the statistical models are inadequate. To address this problem, we propose a cost‐effective, easy to replicate Q‐ band beacon receiver to leverage the Alphasat propagation campaign. We present a step‐by‐step implementation process of the receiver's fundamental part, a Low‐Noise Block, which translates input 39.402 GHz signal into 162 MHz output signal with a conversion gain of 52 dB. The receiver furthermore utilizes software define radio for signal processing and other data manipulation. Here, we describe the receiver implementations in great details, supplementing the crucial parts with laboratory validation results. Finally, we show 2 example datasets, showing usual data obtained during heavy showers and on a quiet day, respectively.     

Preparation of novel fibre–silica–Ag composites: the influence of fibre structure on sorption capacity and antimicrobial activity

Novel fibre–silica–Ag composites with biocidal activity were successfully produced by chemical modifying cotton (CO), wool (WO), silk (SE), polyamide (PA) and polyester (PES) fabrics and CO/PES and WO/PES fabric blends. A silica–Ag coating was prepared using a two-step procedure that included the creation of a silica matrix on the fibre surface via the application of an inorganic–organic hybrid sol–gel precursor [reactive binder (RB)] using a pad-dry-cure method, followed by the in situ synthesis of AgCl particles within the RB-treated fibres from solutions of 0.10 mM and 0.50 mM AgNO₃ and NaCl. The presence of the coating on the fibres was verified by scanning electron microscopy and energy-dispersive X-ray spectroscopy. The bulk concentration of Ag in the coated fibres was determined using inductively coupled plasma mass spectroscopy. The antimicrobial activity was determined for the bacteria Escherichia coli and Staphylococcus aureus and the fungus Aspergillus niger. The results show that the chemical and morphological structures of the fibres directly influenced their absorptivity and affinity for the Ag compound particles. As the amorphous molecular structure of the fibres and the amount of functional groups available as binding sites for Ag⁺ were increased, both the silver solution uptake and the concentration of the absorbed Ag compound particles increased. The chemical binding of Ag to the fibres significantly reduced the effectiveness of the antimicrobial activity of the Ag compound particles. Accordingly, an increase in the concentration of absorbed Ag was required to achieve a biocidal effect.     

Quaternary Ti–20Nb–10Zr–5Ta alloy during immersion in simulated physiological solutions: formation of layers,dissolution and biocompatibility

Samples of the quaternary Ti–20Nb–10Zr–5Ta alloy were immersed in Hanks’ simulated physiological solution and in minimum essential medium (MEM) for 25 days. Samples of Ti metal served as controls. During immersion, the concentration of ions dissolved in MEM was measured by inductively coupled plasma mass spectrometry, while at the end of the experiment the composition of the surface layers was analyzed by X-ray photoelectron spectroscopy, and their morphology by scanning electron microscopy equipped for chemical analysis. The surface layer formed during immersion was comprised primarily of TiO₂ but contained oxides of alloying elements as well. The degree of oxidation differed for different metal cations; while titanium achieved the highest valency, tantalum remained as the metal or is oxidized to its sub-oxides. Calcium phosphate was formed in both solutions, while formation of organic-related species was observed only in MEM. Dissolution of titanium ions was similar for metal and alloy. Among alloying elements, zirconium dissolved in the largest quantity. The long-term effects of alloy implanted in the recipient’s body were investigated in MEM, using two types of human cells—an osteoblast-like cell line and immortalized pulmonary fibroblasts. The in vitro biocompatibility of the quaternary alloy was similar to that of titanium, since no detrimental effects on cell survival, induction of apoptosis, delay of growth, or change in alkaline phosphatase activity were observed on incubation in MEM.     

Cu-7Cr-0.1Ag Microcomposites Optimized for High Strength and High Condutivity

This paper (i) investigated how the microstructure, conductivity, and mechanical properties of Cu-7Cr-0.1Ag microcomposites were changed by cold drawing and subsequent heat treatment, and (ii) produced the Cu-7Cr-0.1Ag microcomposite with an optimum combination of strength and conductivity. The figure of merit Z (combining strength and conductivity) of the Cu-7Cr-0.1Ag microcomposite was larger than that of the microcomposite without silver for each heat treatment. The value of Z of the Cu-7Cr-0.1Ag microcomposite was a maximum after heat treatment for 1 h at 600 °C, indicating that this was the optimum intermediate heat treatment. The following combinations of conductivity, strength and ductility (measured as elongation to fracture) were obtained by the Cu-7Cr-0.1Ag microcomposite with η = 8: (i) 77.9% IACS (International Annealed Copper Standard), 920 MPa and 3.1%; (ii) 79.3% IACS, 880 MPa and 3.3%; and (iii) 79.9% IACS, 798 MPa and 3.5%. These values for the Cu-7Cr-0.1Ag microcomposite were larger than those of the Cu-7Cr microcomposite.     

An automated shotgun lipidomics platform for high throughput,comprehensive, and quantitative analysis of blood plasma intact lipids

Blood plasma has gained protagonism in lipidomics studies due to its availability, uncomplicated collection and preparation, and informative readout of physiological status. At the same time, it is also technically challenging to analyze due to its complex lipid composition affected by many factors, which can hamper the throughput and/or lipidomics coverage. To tackle these issues, we developed a comprehensive, high throughput, and quantitative mass spectrometry‐based shotgun lipidomics platform for blood plasma lipid analyses. The main hallmarks of this technology are (i) it is comprehensive, covering 22 quantifiable different lipid classes encompassing more than 200 lipid species; (ii) it is amenable to high‐throughput, with less than 5 min acquisition time allowing the complete analysis of 200 plasma samples per day; (iii) it achieves absolute quantification, by inclusion of internal standards for every lipid class measured; (iv) it is highly reproducible, achieving an average coefficient of variation of <10% (intra‐day), approx. 10% (inter‐day), and approx. 15% (inter‐site) for most lipid species; (v) it is easily transferable allowing the direct comparison of data acquired in different sites. Moreover, we thoroughly assessed the influence of blood stabilization with different anticoagulants and freeze‐thaw cycles to exclude artifacts generated by sample preparation. Practical applications: This shotgun lipidomics platform can be implemented in different laboratories without compromising reproducibility, allowing multi‐site studies and inter‐laboratory comparisons. This possibility combined with the high‐throughput, broad lipidomic coverage and absolute quantification are important aspects for clinical applications and biomarker research. This MS‐based automated shotgun lipidomics platform for comprehensive analysis of the blood plasma lipidome (covering 22 lipid classes) achieves high inter‐day and inter‐site reproducibility and accuracy and enables unprecedented large scale lipidomics studies.     

The Absence of the AtSYT1 Function Elevates the Adverse Effect of Salt Stress on Photosynthesis in Arabidopsis

Arabidopsis thaliana SYNAPTOTAGMIN 1 (AtSYT1) was shown to be involved in responses to different environmental and biotic stresses. We investigated gas exchange and chlorophyll a fluorescence in Arabidopsis wild-type (WT, ecotype Col-0) and atsyt1 mutant plants irrigated for 48 h with 150 mM NaCl. We found that salt stress significantly decreases net photosynthetic assimilation, effective photochemical quantum yield of photosystem II (Φ_PSII), stomatal conductance and transpiration rate in both genotypes. Salt stress has a more severe impact on atsyt1 plants with increasing effect at higher illumination. Dark respiration, photochemical quenching (qP), non-photochemical quenching and Φ_PSII measured at 750 µmol m⁻² s⁻¹ photosynthetic photon flux density were significantly affected by salt in both genotypes. However, differences between mutant and WT plants were recorded only for qP and Φ_PSII. Decreased photosynthetic efficiency in atsyt1 under salt stress was accompanied by reduced chlorophyll and carotenoid and increased flavonol content in atsyt1 leaves. No differences in the abundance of key proteins participating in photosynthesis (except PsaC and PsbQ) and chlorophyll biosynthesis were found regardless of genotype or salt treatment. Microscopic analysis showed that irrigating plants with salt caused a partial closure of the stomata, and this effect was more pronounced in the mutant than in WT plants. The localization pattern of AtSYT1 was also altered by salt stress.     

Na+-Coupled Respiration and Reshaping of Extracellular Polysaccharide Layer Counteract Monensin-Induced Cation Permeability in Prevotella bryantii B14

Monensin is an ionophore for monovalent cations, which is frequently used to prevent ketosis and to enhance performance in dairy cows. Studies have shown the rumen bacteria Prevotella bryantii B₁4 being less affected by monensin. The present study aimed to reveal more information about the respective molecular mechanisms in P. bryantii, as there is still a lack of knowledge about defense mechanisms against monensin. Cell growth experiments applying increasing concentrations of monensin and incubations up to 72 h were done. Harvested cells were used for label-free quantitative proteomics, enzyme activity measurements, quantification of intracellular sodium and extracellular glucose concentrations and fluorescence microscopy. Our findings confirmed an active cell growth and fermentation activity of P. bryantii B₁4 despite monensin concentrations up to 60 µM. An elevated abundance and activity of the Na⁺-translocating NADH:quinone oxidoreductase counteracted sodium influx caused by monensin. Cell membranes and extracellular polysaccharides were highly influenced by monensin indicated by a reduced number of outer membrane proteins, an increased number of certain glucoside hydrolases and an elevated concentration of extracellular glucose. Thus, a reconstruction of extracellular polysaccharides in P. bryantii in response to monensin is proposed, which is expected to have a negative impact on the substrate binding capacities of this rumen bacterium.