Modeling of a single-cycle current generator while forming a quasi-sinusoidal current

The paper presents the results of investigation of the influence of the output voltage magnitude on the operating frequency of the switch in the single-cycle quasi-sinusoidal current generator circuit. Analytical expressions for calculating the time parameters for transients in the circuit under given assumptions have been obtained. The results presented in the paper can be used in the design of converters of this type

UK–Russia Researcher Links Workshop: extracellular vesicles – mechanisms of biogenesis and roles in disease pathogenesis, M.V. Lomonosov Moscow State University, Moscow, Russia, 1–5 March 2015

The UK–Russia extracellular vesicles (EVs) workshop was held at the Medical Center of the M.V. Lomonosov Moscow State University, Moscow, Russia, with 56 attendees from UK and Russian universities and research institutes. The program consisted of 6 research sessions and was focused on studies of EVs isolated from in vitro model systems or biological fluids, including blood and urine. The multidisciplinary program included presentations on mechanisms of EV biogenesis, the role of EVs in disease pathogenesis, the diagnostic value of EVs, including their quantitation and cargo load, as well as the clinical use of EVs in regenerative medicine. Methodological challenges imposed by the nanoscale size of EVs as well as targeted delivery approaches for therapeutics were considered in a separate session on technologies. The main aim of the workshop was to overview challenges confronting EV researchers and to facilitate knowledge exchange between researchers with different backgrounds and skills. Given the lack of definitive EV nomenclature, specific terms (exosomes or microvesicles) were only applied in the meeting report to studies that carried out full EV characterization, including differential ultracentrifugation isolation approaches, comprehensive protein marker characterization, and single vesicle analysis (electron microscopy and nanoparticle analysis), to ascertain EV size and morphology following the International Society for Extracellular Vesicles standardization recommendations (1,2). In studies where characterization was not conclusive, the term EV is used

Investigating volatile compounds in the Bacteroides secretome

Microorganisms and their hosts communicate with each other by secreting numerous components. This cross-kingdom cell-to-cell signaling involves proteins and small molecules, such as metabolites. These compounds can be secreted across the membrane via numerous transporters and may also be packaged in outer membrane vesicles (OMVs). Among the secreted components, volatile compounds (VOCs) are of particular interest, including butyrate and propionate, which have proven effects on intestinal, immune, and stem cells. Besides short fatty acids, other groups of volatile compounds can be either freely secreted or contained in OMVs. As vesicles might extend their activity far beyond the gastrointestinal tract, study of their cargo, including VOCs, is even more pertinent. This paper is devoted to the VOCs secretome of the Bacteroides genus. Although these bacteria are highly presented in the intestinal microbiota and are known to influence human physiology, their volatile secretome has been studied relatively poorly. The 16 most well-represented Bacteroides species were cultivated; their OMVs were isolated and characterized by NTA and TEM to determine particle morphology and their concentration. In order to analyze the VOCs secretome, we propose a headspace extraction with GC–MS analysis as a new tool for sample preparation and analysis of volatile compounds in culture media and isolated bacterial OMVs. A wide range of released VOCs, both previously characterized and newly described, have been revealed in media after cultivation. We identified more than 60 components of the volatile metabolome in bacterial media, including fatty acids, amino acids, and phenol derivatives, aldehydes and other components. We found active butyrate and indol producers among the analyzed Bacteroides species. For a number of Bacteroides species, OMVs have been isolated and characterized here for the first time as well as volatile compounds analysis in OMVs. We observed a completely different distribution of VOC in vesicles compared to the bacterial media for all analyzed Bacteroides species, including almost complete absence of fatty acids in vesicles. This article provides a comprehensive analysis of the VOCs secreted by Bacteroides species and explores new perspectives in the study of bacterial secretomes in relation the intercellular communication

Comparative Study of Non-Enveloped Icosahedral Viruses Size.

Now, as before, transmission electron microscopy (TEM) is a widely used technique for the determination of virions size. In some studies, dynamic light scattering (DLS) has also been applied for this purpose. Data obtained by different authors and using different methods could vary significantly. The process of TEM sample preparation involves drying on the substrate, which can cause virions to undergo morphology changes. Therefore, other techniques should be used for measurements of virions size in liquid, (i.e. under conditions closer to native). DLS and nanoparticle tracking analysis (NTA) provide supplementary data about the virions hydrodynamic diameter and aggregation state in liquid. In contrast to DLS, NTA data have a higher resolution and also are less sensitive to minor admixtures. In the present work, the size of non-enveloped icosahedral viruses of different nature was analyzed by TEM, DLS and NTA: the viruses used were the encephalomyocarditis virus (animal virus), and cauliflower mosaic virus, brome mosaic virus and bean mild mosaic virus (plant viruses). The same, freshly purified, samples of each virus were used for analysis using the different techniques. The results were compared with earlier published data and description databases. DLS data about the hydrodynamic diameter of bean mild mosaic virus, and NTA data for all examined viruses, were obtained for the first time. For all virus samples, the values of size obtained by TEM were less than virions sizes determined by DLS and NTA. The contribution of the electrical double layer (EDL) in virions hydrodynamic diameter was evaluated. DLS and NTA data adjusted for EDL thickness were in better agreement with TEM results

Adsorption of extracellular vesicles onto the tube walls during storage in solution.

Short term storage of extracellular vesicle (EV) solutions at +4°C is a common practice, but the stability of EVs during this procedure has not been fully understood yet. Using nanoparticle tracking analysis, we have shown that EVs isolated from the conditioned medium of HT-29 cells exhibit a pronounced concentration decrease when stored in PBS in ordinary polypropylene tubes within the range of (0.5-2.1) × 1010 particles/ml. EV losses reach 51±3% for 0.5 ml of EVs in Eppendorf 2 ml tube at 48 hours of storage at +4°C. Around 2/3 of the observed losses have been attributed to the adsorption of vesicles onto tube walls. This result shows that the lower part (up to at least 2 × 1010 particles/ml) of the practically relevant concentration range for purified EVs is prone to adsorption losses at +4°C. Total particle losses could be reduced to 18-21% at 48 hours by using either Eppendorf Protein LoBind tubes or ordinary tubes with the surface blocked with bovine serum albumin or EVs. Reduction of losses to 15% has been shown for isolated EVs dissolved in the supernatant after 100 000 g centrifugation as a model of conditioned medium. Also, a previously unknown feature of diffusion-controlled adsorption was revealed for EVs. In addition to the decrease in particle count, this process causes the predominant losses of smaller particles

Highly Sensitive Measurement of Horseradish Peroxidase Using Surface-Enhanced Raman Scattering of 2,3-Diaminophenazine

The development of various enzyme-linked immunosorbent assays (ELISAs) coupled with surface-enhanced Raman scattering (SERS) detection is a growing area in analytical chemistry due to their potentially high sensitivity. A SERS-based ELISA with horseradish peroxidase (HRP) as an enzymatic label, an o-phenylenediamine (oPD) substrate, and a 2,3-diaminophenazine (DAP) enzymatic product was one of the first examples of such a system. However, the full capabilities of this long-known approach have yet to be revealed. The current study addresses a previously unrecognized problem of SERS detection stage performance. Using silver nanoparticles and model mixtures of oPD and DAP, the effects of the pH, the concentration of the aggregating agent, and the particle surface chloride stabilizer were extensively evaluated. At the optimal mildly acidic pH of 3, a 0.93 to 1 M citrate buffer, and AgNPs stabilized with 20 mM chloride, a two orders of magnitude advantage in the limits of detection (LODs) for SERS compared to colorimetry was demonstrated for both DAP and HRP. The resulting LOD for HRP of 0.067 pmol/L (1.3 amol per assay) underscores that the developed approach is a highly sensitive technique. We suppose that this improved detection system could become a useful tool for the development of SERS-based ELISA protocols

Analysis of virions hydrodynamic size for CaMV (A), EMCV (B), BMV (C) and BMMV (D) using nanoparticle tracking analysis.

<p>Concentration ×10¹² particles per ml (<i>y</i>-axis), diameter in nm (<i>x</i>-axis).</p

Comparative characteristics of the sizes of icosahedral virions using TEM and techniques in liquid (DLS and NTA), considering the contribution of EDL.

<p>Comparative characteristics of the sizes of icosahedral virions using TEM and techniques in liquid (DLS and NTA), considering the contribution of EDL.</p

Transmission electron micrographs of virions CaMV (A), EMCV (B), BMV (C), BMMV (D).

<p>Samples were stained with 2% uranyl acetate. Bars, 100 nm.</p

Analysis of EMCV samples after 2 weeks of storage by NTA (A) and TEM (B).

<p>(A) − Concentration ×10¹² particles per ml (<i>y</i>-axis), hydrodynamic diameter in nm (<i>x</i>-axis).</p