5,812 research outputs found
Block by block: developments in NMR methodology
Due to its high information content and a non-destructive nature, NMR is a versatile and widely used analytical technique. A large variety of different NMR experiments exist, each providing specific information about the system being studied. Over the years a number of building blocks of NMR experiments have been designed, with new still appearing. When combined they can address issues such as spectral complexity, low sample concentrations or signal overlap. This thesis presents work in which several NMR building blocks have been modified and combined creatively to enhance their performance and to produce new, powerful NMR experiments.
The first building block to be utilised is DISPEL (Destruction of Interfering Satellites by Perfect Echo Low-pass filtration),1 a pulse sequence element that suppresses one-bond 13C satellites in 1D 1H spectra. So far it has been utilised to declutter spectra and identify low concentration impurities in samples. In our work, the DISPEL pulse sequence has been combined with a 2D TOCSY experiment to enable the removal of one-bond 13C satellites in 2D correlation spectra. Through comparisons of four DISPEL-TOCSY spectra that can be obtained by varying when the DISPEL elements are active, it was possible to obtain information about site-specific 13C enrichment within molecules. This work has been applied to a sample of partially 13C-labeled amino acids obtained by a digest of E. coli proteins prepared by using 13C-labeled glucose as the carbon source. The DISPEL pulse sequence has also been combined with the JRES experiment aiding the identification of individual peaks and their multiplicity. This work widens the potential of NMR in studies of metabolic pathways.
The second building block explored is the SHARPER (Sensitive, Homogeneous And Resolved PEaks in Real time)2 acquisition technique. This “pure-shift” method removes all heteronuclear and homonuclear couplings from a single signal, reducing the peak to a single singlet with a greatly increased signal to noise ratio, in part due to its inherent capability to compensate for magnetic field inhomogeneity. The original SHARPER pulse sequence was adapted for use on benchtop spectrometers accounting for varying levels of spectrometer hardware and sample requirements. Although equally applicable to high field instruments, this work is particularly beneficial to lower sensitivity, lower magnetic field homogeneity benchtop systems, e. g. for the purpose of reaction monitoring.
By shortening the spin-echo intervals and using non-selective pulses, the SHARPER pulse sequence was modified to remove chemical shift dispersion of all, or a selected group of 1H resonances. This produced a possibility of collapsing an entire spectrum into a single intense and narrow signal that can act as a reporter of molecular properties of studied compounds.
This modification was used to design a SHARPER-DOSY (Diffusion-Ordered SpectroscopY) pulse sequence for the measurement of diffusion coefficients of pure compounds. When applied to small and medium size molecules, the sensitivity of this experiment is 10 to 100-fold higher than that of the original DOSY experiment. It was demonstrated that using cryogenically cooled probes on high-field NMR spectrometers, μM sample concentrations, prepared using as little as 1 μg of a compound, can yield a diffusion coefficient in several minutes.
Finally, SHARPER has been combined with a chemical-shift-selective-filter (CSSF), a technique that enables selective excitation of a single multiplet from a heavily overlapped spectral region, with chemical shift differences as small as 1-2 Hz. The CSSF building block was combined with the DOSY experiment to produce a CSSF-SHARPER-DOSY experiment that enables the determination of diffusion coefficients from highly overlapped spectra
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Liver-directed gene targeting as a potential therapy for Fabry Disease
Fabry disease (FD) is an X-linked inherited, lysosomal storage disorder caused by mutations in the Alpha Galactosidase-A (GLA) gene. This gene encodes for the GLA enzyme which is responsible for the catabolism of glycosphingolipids like globotriaosylceramide (Gb3). Accumulation of Gb3 in lysosomes results in systemic clinical manifestations and reduced lifespan. Enzyme replacement therapy (ERT) and chaperone therapy are the available treatments for FD however, noncurative and with limitations.
We developed a potential therapeutic approach based on the permanent genetic modification of hepatocytes to express the GLA enzyme by targeting the albumin locus in vivo. To model late-onset and early-onset FD, we treated juvenile (P30) and neonatal (P5) Fabry mice with an AAV8 donor vector containing mAlb homology arms and a codon-optimized version of the human GLA cDNA. This treatment was coupled with the AAV-mediated delivery of the CRISPR/SaCas9 platform to increase targeting efficiency. Treatment of juvenile Fabry mice (donor, 3.0E13 vg/kg; SaCas9, 6.0E12 vg/kg) resulted in elevated GLA enzyme activity which was stable till the termination of the experiment at 5 months of age, accompanied by a 70-80% reduction in Lyso-Gb3 accumulation in liver, kidneys and heart, compared to untreated mutant mice. To increase the safety of the procedure, concerns related to the use of programmable nucleases were avoided by applying a nuclease-free approach. Juvenile animals were treated only with the donor vector (3.0E13 vg/kg), coupled with the treatment with fludarabine, which enhances the gene targeting rate. This nuclease-free strategy resulted in increased plasma GLA activity compared to donor-only treated mice, accompanied by 80-95% of lyso-Gb3 clearance.
When we treated neonatal P5 Fabry mice with donor and SaCas9 AAVs, the treatment was significantly more efficient than in juvenile animals due to the increased targeting rate observed in proliferating hepatocytes present in a growing liver. In fact, we were able to completely clear lyso-Gb3 in plasma and in the different organs with the highest dose of 3.0E14 vg/kg of donor vector, while a dose of 3.0E13 vg/kg resulted in a reduction of 95-98% in plasma and target organs.
A dose escalation study with AAV-mediated episomal gene therapy was also done as a proof-of-principle in juvenile Faby KO mice using a strong liver-specific promoter and the human codon-optimized GLA transgene. Animals treated at 3.0E12 vg/kg and higher doses were able to reduce substrate accumulation by 98-100% in plasma and target tissues. Treatment with the lowest dose of the AAV vector (3.0E11vg/kg) resulted in the clearance of 85-95% lyso-Gb3 in the bloodstream and tissues proving the efficacy of the treatment for late-onset FD. ERT-treated animals were considered for comparative evaluation of the treatment.
This data is inclined towards a promising one-shot therapy using an AAV-based integrative gene-editing approach for early-onset and AAV-based episomal gene therapy for late-onset FD to revert the phenotype irrespective of the Fabry disease-causing mutation. To test the translational applicability of this integrative strategy, AAV-LK03 vectors containing human ALB homology arms have been tested in human liver cell lines and will be validated in primary cultures of human hepatocytes, and in humanized mice to generate consistent preclinical support.</br
Long-Molecule Assessment of Ribosomal DNA and RNA
The genes encoding ribosomal RNA and their transcriptional products are essential for life, however, remain poorly understood. Even with the advent of long-range sequencing methodologies, rDNA loci are difficult to study and remain obscure, prompting the consideration of alternative methods to probing this critical region of the genome. The research outlined in this thesis utilises molecular combing, a fibre stretching technique, to isolate DNA molecules measuring more than 5 Mbp in length. The capture of DNA molecules of this size should assist in exploring the architecture of entire rDNA clusters at the single-molecule level. Combining molecular combing with SNP targeting probes, this study aims to distinguish and assess the arrangement of rDNA promoter variants which have been shown to exhibit dramatically different environmental sensitivity. Additionally, through the application of Oxford Nanopore Technologies direct RNA sequencing, the work here has demonstrated the capture of near full-length rRNA primary transcripts, which will allow for assessing post-transcriptional modification across the length of multiple coding subunits within a single molecule, for the first time. Furthermore, an exploration of RNA modification profiles across sample types representative of different developmental stages has been conducted. This study predicts many sites to be differentially modified across these different developmental conditions, several of which are known to be important for, if not crucial in ribosome biogenesis and function. The work outlined in this thesis provides a framework for future studies to conduct long-molecule, genetic, and epitranscriptome profiling of this vital region of the genome, and its dynamic response to a changing environment
LIPIcs, Volume 251, ITCS 2023, Complete Volume
LIPIcs, Volume 251, ITCS 2023, Complete Volum
Effects of municipal smoke-free ordinances on secondhand smoke exposure in the Republic of Korea
ObjectiveTo reduce premature deaths due to secondhand smoke (SHS) exposure among non-smokers, the Republic of Korea (ROK) adopted changes to the National Health Promotion Act, which allowed local governments to enact municipal ordinances to strengthen their authority to designate smoke-free areas and levy penalty fines. In this study, we examined national trends in SHS exposure after the introduction of these municipal ordinances at the city level in 2010.MethodsWe used interrupted time series analysis to assess whether the trends of SHS exposure in the workplace and at home, and the primary cigarette smoking rate changed following the policy adjustment in the national legislation in ROK. Population-standardized data for selected variables were retrieved from a nationally representative survey dataset and used to study the policy action’s effectiveness.ResultsFollowing the change in the legislation, SHS exposure in the workplace reversed course from an increasing (18% per year) trend prior to the introduction of these smoke-free ordinances to a decreasing (−10% per year) trend after adoption and enforcement of these laws (β2 = 0.18, p-value = 0.07; β3 = −0.10, p-value = 0.02). SHS exposure at home (β2 = 0.10, p-value = 0.09; β3 = −0.03, p-value = 0.14) and the primary cigarette smoking rate (β2 = 0.03, p-value = 0.10; β3 = 0.008, p-value = 0.15) showed no significant changes in the sampled period. Although analyses stratified by sex showed that the allowance of municipal ordinances resulted in reduced SHS exposure in the workplace for both males and females, they did not affect the primary cigarette smoking rate as much, especially among females.ConclusionStrengthening the role of local governments by giving them the authority to enact and enforce penalties on SHS exposure violation helped ROK to reduce SHS exposure in the workplace. However, smoking behaviors and related activities seemed to shift to less restrictive areas such as on the streets and in apartment hallways, negating some of the effects due to these ordinances. Future studies should investigate how smoke-free policies beyond public places can further reduce the SHS exposure in ROK
VNHSGE: VietNamese High School Graduation Examination Dataset for Large Language Models
The VNHSGE (VietNamese High School Graduation Examination) dataset, developed
exclusively for evaluating large language models (LLMs), is introduced in this
article. The dataset, which covers nine subjects, was generated from the
Vietnamese National High School Graduation Examination and comparable tests.
300 literary essays have been included, and there are over 19,000
multiple-choice questions on a range of topics. The dataset assesses LLMs in
multitasking situations such as question answering, text generation, reading
comprehension, visual question answering, and more by including both textual
data and accompanying images. Using ChatGPT and BingChat, we evaluated LLMs on
the VNHSGE dataset and contrasted their performance with that of Vietnamese
students to see how well they performed. The results show that ChatGPT and
BingChat both perform at a human level in a number of areas, including
literature, English, history, geography, and civics education. They still have
space to grow, though, especially in the areas of mathematics, physics,
chemistry, and biology. The VNHSGE dataset seeks to provide an adequate
benchmark for assessing the abilities of LLMs with its wide-ranging coverage
and variety of activities. We intend to promote future developments in the
creation of LLMs by making this dataset available to the scientific community,
especially in resolving LLMs' limits in disciplines involving mathematics and
the natural sciences.Comment: 74 pages, 44 figure
Proceedings of SIRM 2023 - The 15th European Conference on Rotordynamics
It was our great honor and pleasure to host the SIRM Conference after 2003 and 2011 for the third time in Darmstadt. Rotordynamics covers a huge variety of different applications and challenges which are all in the scope of this conference. The conference was opened with a keynote lecture given by Rainer Nordmann, one of the three founders of SIRM “Schwingungen in rotierenden Maschinen”. In total 53 papers passed our strict review process and were presented. This impressively shows that rotordynamics is relevant as ever. These contributions cover a very wide spectrum of session topics: fluid bearings and seals; air foil bearings; magnetic bearings; rotor blade interaction; rotor fluid interactions; unbalance and balancing; vibrations in turbomachines; vibration control; instability; electrical machines; monitoring, identification and diagnosis; advanced numerical tools and nonlinearities as well as general rotordynamics. The international character of the conference has been significantly enhanced by the Scientific Board since the 14th SIRM resulting on one hand in an expanded Scientific Committee which meanwhile consists of 31 members from 13 different European countries and on the other hand in the new name “European Conference on Rotordynamics”. This new international profile has also been
emphasized by participants of the 15th SIRM coming from 17 different countries out of three continents. We experienced a vital discussion and dialogue between industry and academia at the conference where roughly one third of the papers were presented by industry and two thirds by academia being an excellent basis to follow a bidirectional transfer what we call xchange at Technical University of Darmstadt. At this point we also want to give our special thanks to the eleven industry sponsors for their great support of the conference. On behalf of the Darmstadt Local Committee I welcome you to read the papers of the 15th SIRM giving you further insight into the topics and presentations
SiPM detector timing response study for the electron-ion collider
The Electron-Ion Collider (EIC) will be a new high luminosity large-scale and high polarization collider designed to investigate the QCD dynamics in the nucleons with
unprecedented precision. It is planned to be built at the Brookhaven National Lab in the US.
Through a dRICH prototype, the performance of Silicon PhotoMultipliers (SiPM), the
baseline photo-sensor candidate for the dRICH was tested.
The employed SiPM readout electronics chip, ALCOR, provides
the time-of-hit measurement through the rollover, coarse and fine time contributions.
In this dissertation, a study on the refinement of the Time Resolution of the Reference Timing system (owing to the fine time correction) is presented.
The corrections applied in order to improve the value of the system Time Resolution
is based on parameters obtained from the measured fine time component of the registered time coincidence signals.
The performance of the calibration procedure described, several checks were performed on dedicated channels.
The results show that it represents an accurate approximation for the correction of 90% of the analysed data.
The performance of the studied SiPMs displayed satisfying results in both
applications - the Imaging SiPMs were successful in registering the Cherenkov light signal and the Timing SiPMs provided a Reference Time value which allowed to correctly
track the signal time-of-hits.
The Reference Timing system was calibrated to provide a measured Time Resolution of 135 ± 2 ps.
A preliminary study of the Imaging sensor Time Resolution, which for was calculated to be for a single photoelectron within approximately 500 ps, indicates that the value of the Timing system Time Resolution is adequate for the framework.
Note that although these preliminary Time Resolution illustrate
satisfactory results, they do not include corrections for effects such as time walk, time over threshold or low sensor bias voltage working conditions, which would presumably further improve
the results
Advances and Applications of DSmT for Information Fusion. Collected Works, Volume 5
This fifth volume on Advances and Applications of DSmT for Information Fusion collects theoretical and applied contributions of researchers working in different fields of applications and in mathematics, and is available in open-access. The collected contributions of this volume have either been published or presented after disseminating the fourth volume in 2015 in international conferences, seminars, workshops and journals, or they are new. The contributions of each part of this volume are chronologically ordered.
First Part of this book presents some theoretical advances on DSmT, dealing mainly with modified Proportional Conflict Redistribution Rules (PCR) of combination with degree of intersection, coarsening techniques, interval calculus for PCR thanks to set inversion via interval analysis (SIVIA), rough set classifiers, canonical decomposition of dichotomous belief functions, fast PCR fusion, fast inter-criteria analysis with PCR, and improved PCR5 and PCR6 rules preserving the (quasi-)neutrality of (quasi-)vacuous belief assignment in the fusion of sources of evidence with their Matlab codes.
Because more applications of DSmT have emerged in the past years since the apparition of the fourth book of DSmT in 2015, the second part of this volume is about selected applications of DSmT mainly in building change detection, object recognition, quality of data association in tracking, perception in robotics, risk assessment for torrent protection and multi-criteria decision-making, multi-modal image fusion, coarsening techniques, recommender system, levee characterization and assessment, human heading perception, trust assessment, robotics, biometrics, failure detection, GPS systems, inter-criteria analysis, group decision, human activity recognition, storm prediction, data association for autonomous vehicles, identification of maritime vessels, fusion of support vector machines (SVM), Silx-Furtif RUST code library for information fusion including PCR rules, and network for ship classification.
Finally, the third part presents interesting contributions related to belief functions in general published or presented along the years since 2015. These contributions are related with decision-making under uncertainty, belief approximations, probability transformations, new distances between belief functions, non-classical multi-criteria decision-making problems with belief functions, generalization of Bayes theorem, image processing, data association, entropy and cross-entropy measures, fuzzy evidence numbers, negator of belief mass, human activity recognition, information fusion for breast cancer therapy, imbalanced data classification, and hybrid techniques mixing deep learning with belief functions as well
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