Exploiting the Symmetry of the Resonator Mode to Enhance PELDOR Sensitivity.

Pulsed electron paramagnetic resonance (EPR) spectroscopy using microwaves at two frequencies can be employed to measure distances between pairs of paramagnets separated by up to 10 nm. The method, combined with site-directed mutagenesis, has become increasingly popular in structural biology for both its selectivity and capability of providing information not accessible through more standard methods such as nuclear magnetic resonance and X-ray crystallography. Despite these advantages, EPR distance measurements suffer from poor sensitivity. One contributing factor is technical: since 65 MHz typically separates the pump and detection frequencies, they cannot both be located at the center of the pseudo-Lorentzian microwave resonance of a single-mode resonator. To maximize the inversion efficiency, the pump pulse is usually placed at the center of the resonance, while the observer frequency is placed in the wing, with consequent reduction in sensitivity. Here, we consider an alternative configuration: by spacing pump and observer frequencies symmetrically with respect to the microwave resonance and by increasing the quality factor, valuable improvement in the signal-to-noise ratio can be obtained

Improving the development, monitoring and reporting of stroke rehabilitation research: consensus-based core recommendations from the Stroke Recovery and Rehabilitation Roundtable (SRRR)

Recent reviews have demonstrated that the quality of stroke rehabilitation research has continued to improve over the last four decades but despite this progress there are still many barriers in moving the field forward. Rigorous development, monitoring and complete reporting of interventions in stroke trials are essential in providing rehabilitation evidence that is robust, meaningful and implementable. An international partnership of stroke rehabilitation experts committed to develop consensus-based core recommendations with a remit of addressing the issues identified as limiting stroke rehabilitation research in the areas of developing, monitoring and reporting stroke rehabilitation interventions. Work exploring each of the three areas took place via multiple teleconferences and a two-day meeting in Philadelphia in May 2016. A total of 15 recommendations were made. To validate the need for the recommendations the group reviewed all stroke rehabilitation trials published in 2015 (n=182 papers). Our review highlighted that the majority of publications did not clearly describe how interventions were developed or monitored during the trial. In particular, under-reporting of the theoretical rationale for the intervention and the components of the intervention calls into question many interventions that have been evaluated for efficacy. More trials were found to have addressed the reporting of interventions recommendations than those related to development or monitoring. Nonetheless the majority of reporting recommendations were still not adequately described. To progress the field of stroke rehabilitation research and to ensure stroke patients receive optimal evidence based clinical care we urge the research community to endorse and adopt our recommendations

Moiré patterns observed in bi layer graphene irradiated with high energetic protons

Customarily, it is likely that irradiated graphene yield indication of per- turbations induced by irradiation. High Resolution Transmission Electron Micros- copy (HRTEM) analysis has been performed on proton irradiated graphene. The analysis indicates the existence of Moiré patterns produced by the rotations induced by the irradiation in between planes. The rotations measured fluctuate between 3 and 5 degrees respectively. These rotations may influence the electronic properties of the material under investigation. In order to explain the observed rotations in between planes, theoretical analysis were performed under the scheme of extended Hückel tight-binding method. Average total energy of the system was careful ana- lyzed throughout the experiment composed of two graphene layers with two carbon vacancies and then the replaced carbons were intercalated in between the two lay- ers. The results obtained indicate that the system remain semi metallic. Moreover, the theoretical results yielded that the 3 degree rotation is favored, although the 5 degree rotation is not discarded. Furthermore, energy bands as well as total and projected DOS were performed in order to provide more information about the electronic changes induced by the rotations applied to the system

Therapeutic limitations in tumor-specific CD8+ memory T cell engraftment

BACKGROUND: Adoptive immunotherapy with cytotoxic T lymphocytes (CTL) represents an alternative approach to treating solid tumors. Ideally, this would confer long-term protection against tumor. We previously demonstrated that in vitro-generated tumor-specific CTL from the ovalbumin (OVA)-specific OT-I T cell receptor transgenic mouse persisted long after adoptive transfer as memory T cells. When recipient mice were challenged with the OVA-expressing E.G7 thymoma, tumor growth was delayed and sometimes prevented. The reasons for therapeutic failures were not clear. METHODS: OT-I CTL were adoptively transferred to C57BL/6 mice 21 – 28 days prior to tumor challenge. At this time, the donor cells had the phenotypical and functional characteristics of memory CD8+ T cells. Recipients which developed tumor despite adoptive immunotherapy were analyzed to evaluate the reason(s) for therapeutic failure. RESULTS: Dose-response studies demonstrated that the degree of tumor protection was directly proportional to the number of OT-I CTL adoptively transferred. At a low dose of OT-I CTL, therapeutic failure was attributed to insufficient numbers of OT-I T cells that persisted in vivo, rather than mechanisms that actively suppressed or anergized the OT-I T cells. In recipients of high numbers of OT-I CTL, the E.G7 tumor that developed was shown to be resistant to fresh OT-I CTL when examined ex vivo. Furthermore, these same tumor cells no longer secreted a detectable level of OVA. In this case, resistance to immunotherapy was secondary to selection of clones of E.G7 that expressed a lower level of tumor antigen. CONCLUSIONS: Memory engraftment with tumor-specific CTL provides long-term protection against tumor. However, there are several limitations to this immunotherapeutic strategy, especially when targeting a single antigen. This study illustrates the importance of administering large numbers of effectors to engraft sufficiently efficacious immunologic memory. It also demonstrates the importance of targeting several antigens when developing vaccine strategies for cancer

Quantification of nitrotyrosine in nitrated proteins

For kinetic studies of protein nitration reactions, we have developed a method for the quantification of nitrotyrosine residues in protein molecules by liquid chromatography coupled to a diode array detector of ultraviolet-visible absorption. Nitrated bovine serum albumin (BSA) and nitrated ovalbumin (OVA) were synthesized and used as standards for the determination of the protein nitration degree (ND), which is defined as the average number of nitrotyrosine residues divided by the total number of tyrosine residues in a protein molecule. The obtained calibration curves of the ratio of chromatographic peak areas of absorbance at 357 and at 280 nm vs. nitration degree are nearly the same for BSA and OVA (relative deviations <5%). They are near-linear at low ND (< 0.1) and can be described by a second-order polynomial fit up to \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$${\hbox{ND}} = 0.5\left( {{R^2} > 0.99} \right)$$\end{document}. A change of chromatographic column led to changes in absolute peak areas but not in the peak area ratios and related calibration functions, which confirms the robustness of the analytical method. First results of laboratory experiments confirm that the method is applicable for the investigation of the reaction kinetics of protein nitration. The main advantage over alternative methods is that nitration degrees can be efficiently determined without hydrolysis or digestion of the investigated protein molecules

Thermal and Sedimentation Stress Are Unlikely Causes of Brown Spot Syndrome in the Coral Reef Sponge, Ianthella basta

Background: Marine diseases are being increasingly linked to anthropogenic factors including global and local stressors. On the Great Barrier Reef, up to 66% of the Ianthella basta population was recently found to be afflicted by a syndrome characterized by brown spot lesions and necrotic tissue.\ud \ud Methodology/Principal Findings: Manipulative experiments were undertaken to ascertain the role of environmental stressors in this syndrome. Specifically, the effects of elevated temperature and sedimentation on sponge health and symbiont stability in I. basta were examined. Neither elevated temperature nor increased sedimentation were responsible for the brown spot lesions, but sponges exposed to 32°C developed substantial discoloration and deterioration of their tissues, resulting in death after eight days and a higher microbial diversity in those samples. No shifts in the microbial community of I. basta were observed across a latitudinal gradient or with increased sedimentation, with three previously described symbionts dominating the community of all sponges (Alphaproteobacteria, Gammaproteobacteria and Thaumarchaea).\ud \ud Conclusions/Significance: Results from this study highlight the stable microbial community of I. basta and indicate that thermal and sedimentation stress are not responsible for the brown spot lesions currently affecting this abundant and ecologically important sponge species

Evaluation of whole genome sequencing for outbreak detection of Salmonella enterica

Salmonella enterica is a common cause of minor and large food borne outbreaks. To achieve successful and nearly 'real-time' monitoring and identification of outbreaks, reliable sub-typing is essential. Whole genome sequencing (WGS) shows great promises for using as a routine epidemiological typing tool. Here we evaluate WGS for typing of S. Typhimurium including different approaches for analyzing and comparing the data. A collection of 34 S. Typhimurium isolates was sequenced. This consisted of 18 isolates from six outbreaks and 16 epidemiologically unrelated background strains. In addition, 8 S. Enteritidis and 5 S. Derby were also sequenced and used for comparison. A number of different bioinformatics approaches were applied on the data; including pan-genome tree, k-mer tree, nucleotide difference tree and SNP tree. The outcome of each approach was evaluated in relation to the association of the isolates to specific outbreaks. The pan-genome tree clustered 65% of the S. Typhimurium isolates according to the pre-defined epidemiology, the k-mer tree 88%, the nucleotide difference tree 100% and the SNP tree 100% of the strains within S. Typhimurium. The resulting outcome of the four phylogenetic analyses were also compared to PFGE revealing that WGS typing achieved the greater performance than the traditional method. In conclusion, for S. Typhimurium, SNP analysis and nucleotide difference approach of WGS data seem to be the superior methods for epidemiological typing compared to other phylogenetic analytic approaches that may be used on WGS. These approaches were also superior to the more classical typing method, PFGE. Our study also indicates that WGS alone is insufficient to determine whether strains are related or un-related to outbreaks. This still requires the combination of epidemiological data and whole genome sequencing results