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1
Fu, Shilin. "Prediction of electrophoretic mobilities in capillary zone electrophoresis". Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1998. http://www.collectionscanada.ca/obj/s4/f2/dsk2/ftp03/MQ31347.pdf.
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FINETTI, CHIARA. "NOVEL FUNCTIONAL HYDROPHILIC POLYMERS AND HYDROGELS FOR MICROANALYTICAL SYSTEMS". Doctoral thesis, Università degli Studi di Milano, 2017. http://hdl.handle.net/2434/473212.
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This thesis focuses on the use of hydrophilic polymers for bioanalytical applications, including several microanalytical techniques encompassing nanotechnology, microarray technology and DNA gel electrophoresis.
The dissertation is divided in two parts, which share the employment of dimethylacrylamide-based copolymers, developed at the laboratory of Analytical Microsystems of the Institute of Chemistry for Molecular Recognition (National Research Council of Italy) where the thesis has been carried out.
PART A introduces a novel approach for surface modification of quantum dots and gold nanoparticles, based on physi-/chemisorption of two different functional dimethylacrylamide copolymers. Beside developing innovative functionalization strategies, the goal is to demonstrate the application of coated nanoparticles in highly sensitive immunoassays based on microarray technology.
PART B of the dissertation presents the results of an activity, conducted in collaboration with the company Agilent Technology (UK), aimed at developing an innovative gel sieving matrix for high performance DNA electrophoresis. We introduce a new hydrogel obtained by cross-linking an alkyne modified polymer with an azide one, exploiting a copper catalysed click chemistry reaction. The alkyne functionalized polymer is based on poly(dimethilacrylamide) and it was obtained by a post-polymerization modification approach from the parent copolymer poly(DMA-NAS-MAPS), extensively used in the first part of this dissertation. The azide polymer is a polyethylenglycol terminated with azide groups at both ends, and is commercially available.
A considerable part of this work is devoted to the optimization of the characteristics of the new hydrogel, in particular to the extension of its shelf-life, an important parameter in view of its industrial application.
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OTTONE, TIZIANA. "La mutazione di tipo A della nucleofosmina nella diagnosi e nel follow-up della leucemia mieloide acuta". Doctoral thesis, Università degli Studi di Roma "Tor Vergata", 2009. http://hdl.handle.net/2108/1135.
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Acute myeloid leukaemias (AML), which account for about 80% of the acute leukaemias in adult, represent a phenotypically and genetically heterogeneous group of clonal haematopoietic diseases in which, failure to differentiate and over proliferation in the stem cell/progenitor compartment, result in accumulation of non-functional myeloblasts. Nucleophosmin-1 (NPM1) mutations is the most frequent gene alteration in AML (30%), and are particularly frequent in AML with normal karyotype (AML-NK) (60%). These alterations have been shown to carry prognostic significance because they seem to identify patients with better response to chemotherapy. As a consequence, the analysis of NPM1 mutational status is now recommended for inclusion in the genetic routine characterization of AML (Gorello P. et al., Leukemia 2006; Ammatuna E. et al., Clin Chem 2005; Nelida I.N. et al., Leukemia 2005; Scholl S. et al., Leuk Res 2007; Lin LI et al., Leukemia 2006).
Finally, given their high prevalence and stability over the course of the disease, NPM1 mutations may serve as an ideal target for minimal residual disease (MRD) assessment, particularly for patients with AML-NK (Gorello P. et al., Leukemia 2006).
In early 2005, Falini et al. (Falini B. et al., New Engl J Med 2005) reported that NPM1 gene mutations, mainly consisting of small nucleotide insertion/deletions, occur frequently in AML and are strongly associated with NK. Over 40 different types of mutations in the NPM1 (NPM1m) locus have been described so far, which result in the formation of different mutant proteins. This alterations consist of the insertion/deletion of short nucleotide stretches (4 or 10 bps) after nucleotide position 956 in NPM1 exon 12 that lead to an ORF shift resulting in different protein variants containig novel C-terminus portions. The most common NPM1 mutation is the type A (NPM1 mut-A), accounting for 75% - 85% of cases, and consist of duplication of a TCTG tetranucleotide at position 956 to 959 of the reference sequence (GenBank accession number NM_002520).
Available methods to detect nucleophosmin mutations are costly, require sophisticated equipment, and are often less sensitive. The aim of this work was to develop two assays that enable rapid and sensitive detection of NPM1 mut-A (Ottone T. et al., Genes Chromosomes Cancer 2009): a fragment analysis method and an RT-ASO-PCR (Reverse Trascriptase Allele Specific Oligonucleotide Polymerase Chain Reaction) method. Furthermore a semi-nested ASO-PCR method was also designed in order to increase the sensitivity of our assay for the monitoring of MRD.
After informed consent, bone marrow and peripheral blood cells were collected at presentation from 107 patients with AML diagnosed at the Institute of Hematology, Department of Biopathology of the University Tor Vergata of Rome. For each patient reverse trascribed cDNA was divided into three aliquotes used for capillary elettrophoresis, sequencing and ASO-PCR. Semi-nested ASO-PCR experiments were carried out using RT-ASO-PCR products. For capillary electrophoresis fluorescent labeled PCR products were used to screen for the presence of NPM1 mutations using the CEQTM 8000 Genetic Analysis Sistem (Beckman Coulter) instrument. Electroferogram showed 348 bp and 352 bp fragment size, corresponding to NPM1wt and NPM1m, respectively. OCI-Aml3 cell line was used as a positive control. The sensitivity of method was tested using the following serial dilutions of the mutated with wild type allele: 0.01, 0.10, 0.25, 0.50, 0.75, 1.00 (mut/wt). To confirm fragment analysis results, 17 NPM1m and 10 NPM1wt samples were sequenced: cDNA were amplified and the products, after purification, were prepared for sequencing using the CEQTM 8000 instrument.
With the aim of analyzing the presence of type A mutation in NPM1 exon 12, an RT-ASO-PCR strategy was used. A forward primer was designed to specifically amplify NPM1 exon 12 only if the mutation A is present; this primer in fact contains an intentional mismatch at third nucleotide from 3’ end to improve specificity. The amplified region includes the insertion of a TCTG tetranucleotide and corresponds to a 320 bp amplified product. As internal PCR control the ABL housekeeping gene was also amplified in each sample.
In order to increase the sensitivity for the assay for MRD monitoring, we set up a semi-nested ASO-PCR; for this purpose an internal forward primer NPM-AN for NPM1 mut-A was designed, corresponding to a 319 bp amplified fragment.
Fragment analysis allowed to identify 17 NPM1m samples. All the NPM1m samples were heterozygous confirming that NPM1m allele is negative dominant. As concerning sensitivity method enabled to detect the NPM1 mutations present in concentration of 10-2.
Sequencing analysis confirmed in all instances NPM1m and NPM1wt cases, showing that 12 samples harboured the most frequent type A mutation, the remaining mutated samples showing type B (3 cases), type D (1 case) and type K mutation (1 case).
RT-ASO-PCR assay confirmed the capillary electrophoresis and sequencing results, showing a concordance in 100% of cases; in fact this method identified the 12 out of 17 NPM1m samples as NPM1 mut-A, with an overall frequency of 70%.
The semi-nested ASO-PCR assay was carried out to increase the sensitivity of this assay for the monitoring of MRD. The sensitivity calculated for both methods, RT-ASO-PCR and semi-nested ASO-PCR, indicated that this assays could detect the NPM1 mut-A present in concentration of 10-2 and 10-5, respectively.
Out of 107 patients, cytogenetic data were available for 99 patients, divided into 2 groups: a group (n=15) consisting of NPM1m and a group (n=84) with NPMwt. Twenty seven out of 99 had normal karyotype. Amongst twenty seven, 13 were NPM1m with an incidence of 48%.
The capillary electrophoresis assay developed in this work proved to be a fast, reproducible and easy method applicable to NPM1m diagnosis, capable to discriminate 1 mutated cell out of 100. RT-ASO-PCR and the more sensitive semi-nested ASO-PCR were developed for specifically detect NPM1mut-A, the most frequent NPM1 mutation. These methods were capable to detect 1 mutated cell out of 100 and out of 100.000, respectively.
Monitoring more patients by these new assays in MRD longitudinal studies could be relevant for better assess molecular remission and the risk of relapse in AML-NK.
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Lisi, Samuele. "Approches innovantes basées sur la Résonance des Plasmons de Surface pour le diagnostic biomoléculaire de la maladie d’Alzheimer". Thesis, Université Grenoble Alpes (ComUE), 2017. http://www.theses.fr/2017GREAV003/document.
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La maladie d’Alzheimer est une pathologie neurodégénérative qui amène à une perte progressive de la mémoire et cause des changements comportementaux. Selon plusieurs théories, le développement de cette maladie est associé à l’accumulation du peptide amyloïde beta et de la protéine tau dans des zones précises du cerveau humain. A l’heure actuelle, les approches thérapeutiques testées sont fondées sur l’hypothèse de la cascade amyloïde, mais les résultats n’ont pas été jugés suffisamment efficaces. Pour augmenter les chances de succès des traitements thérapeutiques existants, de meilleures techniques pour un dépistage précoce de l’Alzheimer semblent nécessaires. De ce fait, dans cette thèse, des stratégies innovantes pour l’analyse d’un des biomarqueurs de la maladie d’Alzheimer sont proposées. En particulier le projet porte sur l’analyse de la protéine tau avec des biocapteurs basés sur la Résonance de Plasmons de Surface (SPR). L’augmentation du niveau de ce biomarqueur dans le Liquide Céphalo-Rachidien (LCR) est déjà indicateur d’un processus de neurodégénérescence. De plus, si la mesure de la protéine tau est combinée à celle d’autres biomarqueurs de la pathologie (i.e. : amyloïde beta), les possibilités de dépistage sont fortement augmentées. Les travaux ont portés sur deux aspects : initialement l’interaction antigène-anticorps a été exploitée pour développer un immunocapteur pour la protéine tau. En utilisant cette technologie, nous avons pu caractériser les paramètres analytiques de l’essai direct (avec un seul anticorps) et ceux de l’essai sandwich (avec deux anticorps complémentaires). Dès ces premières approches, nous avons remarqué le besoin d’augmenter la sensibilité de la méthode SPR développée. En effet la limite de détection pour l’essai sandwich était de l’ordre du nM, alors que les niveaux de tau dans le LCR sont de l’ordre du pM. L’utilisation de nanotechnologies, en particulier des nanotubes de carbone, a permis d’atteindre des niveaux proches du pM, avec de bonnes performances en terme de répétabilité de l’essai.Une approche alternative a été conçue dans la deuxième partie du projet. Elle était consacrée à la sélection d’un aptamère pour la protéine tau, afin d’exploiter les avantages de cette classe de récepteurs par rapport aux anticorps. Pour accomplir cet objectif, deux stratégies de sélection ont été mises en place. Premièrement la sélection traditionnelle (SELEX, Systematic Evolution of Ligands by EXponential enrichment) a été appliquée en utilisant l’Electrophorèse Capillaire (EC) comme moyen de séparation. Bien que de nombreuses conditions aient été modifiées, avec le SELEX traditionnel nous n’avons pas observé une évolution significative de l’affinité entre les séquences d’ADN et la protéine tau. Dans la deuxième approche nous avons utilisé la même méthode de séparation pour mener la sélection à travers l’EC-Non-SELEX. En utilisant cette méthode, où les étapes de PCR étaient réduites, une évolution positive a été observée après seulement trois rounds. En effet cinq séquences parmi celles issues du dernier round ont montré une affinité supérieure pour la cible par rapport à la banque. Néanmoins le nombre de séquences analysées à la fois par SPR et par anisotropie de fluorescence reste extrêmement limité par rapport au pool initial. Même si ceci semble être une limite, ce travail est le premier où les aptamères sont appliqués à l’analyse de la protéine tau. Le potentiel de cette classe de récepteurs reste en grande partie inexploré, ce qui laisse entrevoir des améliorations possibles de l’affinité grâce à de meilleurs processus de sélection et au développement de nouveaux outils bioinformatiques.En conclusion la SPR grâce à ses caractéristiques jouera un rôle fondamental dans les prochaines années pour l’analyse des biomarqueurs et pour le screening de nouvelles molécules, qui seront l’objet de futurs essais cliniques pour limiter l’agrégation de la protéine tauAlzheimer’s disease (AD) is a widespread pathogenic condition causing memory and behavior impairment mostly in elderlies because of the accumulation of amyloid beta peptide and tau protein in human brain. Current therapeutic approaches, based on the amyloid hypothesis, are unable to arrest the progression of the disease, hence early diagnosis is crucial for an effective intervention. Based on the updated criteria for AD probable diagnosis, and considering the limits associated with the actual analytical techniques, my work in this thesis was dedicated to develop novel strategies for AD diagnosis. The whole project focused on the analysis of tau protein by Surface Plasmon Resonance (SPR) biosensing. Such protein is well known for being relevant as neurodegenerative marker. In particular if the measurement of tau is associated with that of the amyloid beta peptide and that of the phosphorylated tau, the clinical specificity of this protein become significant to detect Alzheimer. Two aspects were studied; first of all an immunosensor was developed taking advantage of the well-established antigen-antibody interaction. After characterization of the analytical parameters of the direct assay (with primary antibody), a sandwich assay (using two monoclonal antibodies mapping on different analyte i.e. protein tau epitopes) was developed, allowing very low sensitivity to be obtained in artificial Cerebrospinal Fluid (aCSF). In particular to enhance the analytical signal Carbon Nano Tubes (CNTs) were used. Secondly, the research was focused on the selection of aptamers for tau. To this aim two SELEX (Systematic Evolution of Ligands by EXponential enrichment) methods were compared, both based on Capillary Electrophoresis (CE) for partitioning step of the process. Whether with CE-SELEX (first method), no significant affinity improvement was measured, using the CE-Non-SELEX (second method) affinity of the DNA library for tau protein was consistently improved. After isolation of a limited population of aptamer candidates, five sequences were chosen to be analyzed for their affinity for the target. Fluorescence Anisotropy (FA) measurements and SPR highlight similar behavior for the selected sequences, despite the detection principles of these techniques are significantly different. In conclusion the work highlight versatility of SPR technology used both for quantitative analysis and for new selected aptamers characterization in terms of affinity for the analyte tau. The above mentioned versatility is of great interest in a field such AD, which is rapidly expanding. Lowering the total tau levels has been recently identified as a new goal for therapy. Therefore many drug candidates are likely going to be tested in the near future. SPR technology is already widely used in pharmaceutical industry to investigate novel molecules, since it gives access to a large panel of information. In this panorama aptamer technology may improve the overall quality of the analytical data, allowing better comparison among drug candidates. With respect of these receptors, the thesis opened the door to new studies for DNA aptamers to recognize tau, with considerable advantages in term of the receptor stability. Moreover the whole potential of DNA aptamers selected in this work still remain to be explored. New selection methodologies, combined with fast progression of bioinformatics tools might give rise to affinity improvement, which will lead to sensitivity improvement for tau detection in the next few years
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LUCARELLI, CLAUDIA. "La multiresistenza in Salmonella: caratterizzazione molecolare di un nuovo clone emergente di Salmonella enterica sierotipo Typhimurium". Doctoral thesis, Università degli Studi di Roma "Tor Vergata", 2009. http://hdl.handle.net/2108/1089.
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Salmonella enterica sierotipo Typhimurium (STM) rappresenta la prevalente causa di gastroenterite trasmessa da alimenti in Italia, con la maggior parte degli isolati con resistenza multipla agli antibiotici, principalmente ad ampicillina (A), cloramfenicolo (C), streptomicina (S), sulfamidici (Su) e tetraciclina (T) (ACSSuT).
Un nuovo pattern di resistenza (R-type) ASSuT, mancante della resistenza a C, è recentemente emerso in Italia tra ceppi di STM e della sua variante monofasica, Salmonella enterica subspecie enterica sierotipo S. 4,[5],12:i:– .
L’obiettivo principale di questa tesi è stato la caratterizzazione di ceppi di STM e di S. 4,[5],12:i:– con R-type ASSuT, usando tecniche di tipizzazione molecolari e fenotipiche, quali l’elettroforesi in campo pulsato (PFGE) e la fagotipizzazione, allo scopo di valutare la loro origine clonale e la loro relazione con i ceppi ACSSuT. Usando il database Pulse-Net Europe è stata valutata la presenza di ceppi ASSuT in altre nazioni Europee al fine di allestire una collezione internazionale di ceppi. Questa collezione è stata ulteriormente caratterizzata, identificando i geni di resistenza, investigando la loro localizzazione, e determinando la regione di resistenza.
Sia tra ceppi di STM che di S. 4,[5],12:i:–, il principale profilo di PFGE è rappresentato da STYMXB.0079, mentre i ceppi STM ACSSuT appartengono ai profili STYMXB.0061 e STYMXB.0067. L’analisi dei profili di PFGE con il software Bionumerics ha mostrato che più del 90% dei ceppi ASSuT e ACSSuT appartenevano a due distinti clusters con un’omologia genetica del 73%, dati che dimostrano l’appartenenza dei ceppi ASSuT ad un'unica linea clonale differente da quella dei ceppi ACSSuT.
La maggior parte dei ceppi con profilo ASSuT non erano tipizzabili (DTNT) attraverso la fagotipizzazione o appartenevano al fagotipo U302. Al contrario i ceppi ACSSuT appartenevano principalmente al fagotipo DT104.
Successivamente, nel database Pulse-Net Europe, è stato possibile identificare ceppi ASSuT, sia STM che S. 4,[5],12:i:– isolati in Danimarca ed Inghilterra, con profili di PFGE identici o strettamente correlati a quelli dei ceppi italiani, dati che indicano che il clone ASSuT è presente anche in altri paesi Europei.
Al fine di identificare i geni responsabili della resistenza sono stati selezionati 64 ceppi di STM e S. 4,[5],12:i:– ASSuT, e 11 ceppi di STM con differenti R-type e profili di PFGE, usati come controlli. Tutti i ceppi provenivano da infezioni umane ed erano stati isolati in Italia, Danimarca e Inghilterra.
Tutti i ceppi ASSuT erano positivi per i seguenti geni di resistenza: blaTEM, strA-strB, sul2 and tet(B). Successivi esperimenti di localizzazione hanno dimostrato che i geni di resistenza ASSuT sono localizzati sul cromosoma
Infine, è stata determinata la sequenza completa del cluster di resistenza ASSuT. Questo cluster è composto da due isole di resistenza (RI1 e RI2) separate da DNA cromosomale. In particolare, RI1 è compresa tra due IS26 e contiene deltatnp3R, blaTEM-1, tnpB, seguito da strB, strA, sul2, repC, deltarepA ed un’atra IS26. RI2 è anch’essa compresa tra due IS26, comprendenti deltaIS10L, il gene di resistenza alla tetraciclina, IS1, l’operone per la resistenza al mercurio, il gene yaeA, e un’ ipotetica transposasi (tniAdelta). Entrambe le RIs mostrano il 99% di identità con due regioni adiacenti del plasmide pHCM1, presente nel ceppo di S. Typhi isolato in Vietnam. Le sequenze di inserzione IS26 potrebbero aver avuto un ruolo nella formazione di questo cluster di resistenza, ma quest’ipotesi deve essere ancora verificata.
In conclusione il lavoro di questa tesi indica che i ceppi ASSuT di STM e S. 4,[5],12:i:–, in aumento in Italia, appartengono ad un'unica linea clonale e che i ceppi S. 4,[5],12:i:– circolanti nella nostra nazione, derivano principalmente da questa linea clonale di STM. Inoltre il clone ASSuT è diffuso anche in Danimarca ed Inghilterra. Il pattern di antibiotico resistenza conferito da un’isola di resistenza cromosomale, con un’organizzazione simile ad altri cluster precedentemente descritti, suscita preoccupazione poichè la resistenza può essere mantenuta stabilmente in assenza di pressione selettiva.Salmonella enterica serovar Typhimurium (STM) represents the prevalent cause of foodborne gastroenteritis in Italy with the majority of isolates exhibiting multidrug resistance, mainly to ampicillin (A), chloramphenicol (C), streptomycin (S), sulfonamide (Su) and tetracycline (T) (ACSSuT). However, a new resistance pattern (R-type) ASSuT, lacking resistance to C, has recently emerged in Italy among strains of STM and of its monophasic variant, Salmonella enterica subspecie enterica serovar S. 4,[5],12:i:– . The main objective of this thesis has been the characterization of STM and S. 4,[5],12:i:– strains with R-type ASSuT, using both molecular and phenotypic typing technique, pulsed-field gel electrophoresis (PFGE) and phage typing, in order to evaluate their clonal origin and the relationships with the ACSSuT strains. In addition, by the use of the Pulse-Net database it was evaluated if ASSuT strains were present in other European countries in order to set up an international collection of these strains. This collection has been further characterized with the identification of resistance genes, the investigation of their localization, and determination of the resistance region. Among both the STM and S. 4,[5],12:i:– ASSuT strains, the predominant PFGE profile was STYMXB.0079, while the STM ACSSuT strains belonged to the STYMXB.0061 and STYMXB. 0067. Bionumerics cluster analysis of PFGE profiles showed that more than 90% of ASSuT and ACSSuT resistant strains were included in two distinct clusters with a genetic homology of 73% each other, suggesting that the ASSuT resistant strains belong to a same clonal lineage different from that of the ACSSuT strains. Phage typing showed that both STM and S. 4,[5],12:i:– ASSuT strains were not typeable (DTNT) or U302. A different figure was observed for the ACSSuT strains: the STM strains mostly belonged to DT104. The Pulse-Net Europe database, allowed us to identify ASSuT strains, both STM and S. enterica 4,[5],12:i:–, isolated in Denmark and UK, with the same or very closely related PFGE patterns as the Italian strains, suggesting that the ASSuT clone is circulating in different European countries. The resistance genes were identified in 64 strains of STM and S. enterica 4,[5],12:i:–with ASSuT R-type and in 11 STM strains with different resistance patterns and PFGE profiles as controls. All strains were isolated from human infections in Italy, Denmark and UK. All the ASSuT strains were positive for the following resistance genes: blaTEM, strA-strB, sul2 and tet(B). The control strains showed the same gene pattern, in accordance with their resistance profiles. A variability of the genes conferring resistance to tetracycline was detected. Localization experiments demonstrated that the ASSuT resistance genes are chromosomally located. Finally, the complete sequence of ASSuT resistance cluster was determined. This cluster is composed by two resistance island (RI1 and RI2) divided by chromosomal DNA. In particular, RI1 is comprised between two IS26 and contains deltatnp3R, blaTEM-1, tnpB , followed by strB, strA, sul2, repC, DeltarepA and another IS26. RI2 is bracketed by two IS26, comprising deltaIS10L, tetracycline resistance gene, IS1, the operon for resistance to mercury, yaeA gene, and a putative transposase (tniAdelta). Both this RIs show 99% sequence identity to two adiacent region of pHCM1 plasmid, harbored in S. Typhi isolated in Vietnam. IS26 elements could have played a role in the assembly of this resistance cluster but it will be investigated more in detail. In conclusion the work of this thesis indicates that the tetra-resistant ASSuT strains of STM and S. 4,[5],12:i:–, increasingly isolated in Italy, belong to a same clonal lineage and that the S. 4,[5],12:i:– strains circulating in our country, mainly derive from this STM clonal lineage. ASSuT clone is also circulating in Denmark and United Kingdom. The antimicrobial resistance pattern conferred by a chromosomal island, with an organization similar to previously reported clusters, deserves concern since the resistance could be stably maintained even in the absence of selective pressure.
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Khalifeh, Iman. "Determination of self association constant between bovine insulin molecules by capillary zone electrophoresis". Thesis, Uppsala University, Department of Medical Biochemistry and Microbiology, 2005. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-6155.
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Capillary electrophoresis (CE) is an analytical technique that is very useful for investigating processes that modify the charge and mass of proteins and polypeptide pharmaceuticals. This report explores the ability of CE to determine the aggregation constant between insulin molecules. Bovine insulin is a polypeptide (Mw=5733, pI = 5.3) that has two α-amino groups (Gly and Phe) and one ε–amino group (Lys). Analysis of concentration dependence of electrophoretic mobility of insulin at different conditions yields the association constant for dimerization of insulin. The association constant estimates how tight the peptide molecules are associated. The association constant is a useful factor to evaluate the purity of a peptide or protein sample.
The association reaction of bovine insulin molecules was found to be favoured by temperature. The association constants were 7200 M -1, 8000 M -1, and 36000 M -1 at 15 oC, 25 oC and 35 oC, respectively. The interactions between the peptide molecules increase at higher temperature, resulting in stronger association. The association constant was estimated to be 3000 M -1in the presence of dioxane (5%, w/v %) at 25 oC. However, the interaction sites remain to be explored.
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Vuorensola, Katariina. "Capillary electrophoresis and capillary electrophoresis-mass spectrometry in catecholamine studies". Helsinki : University of Helsinki, 2002. http://ethesis.helsinki.fi/julkaisut/mat/kemia/vk/vuorensola/.
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Renard, Charly. "Nouvelles approches pour la quantification et la réduction de l’adsorption de biomolécules en électrophorèse capillaire : capillaires superhydrophobes et multicouches de polyélectrolytes". Thesis, Montpellier, 2020. http://www.theses.fr/2020MONTS013.
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L’objectif de cette thèse est d’étudier différentes approches pour la modification de la paroi interne des capillaires d’électrophorèse capillaire pour améliorer les efficacités de séparation de biomolécules (peptides et/ou protéines modèles) en milieu acide. Le premier chapitre (hors étude bibliographique) est consacré à l’étude des revêtements superhydrophobes, dans le but d’empêcher l’adsorption des analytes en supprimant toute interaction entre la paroi superhydrophobe et les analytes (absence de mouillage). Un protocole de greffage original a été développé pour obtenir des capillaires superhydrophobes en étudiant l’influence du nombre de couches déposées, la nature du revêtement, et les pressions de remplissage et de rinçage du capillaire lors du dépôt des couches. Des capillaires superhydrophobes ont pu être obtenus, pour la première fois, avec des diamètres internes allant de 50 µm à 180 µm. Les caractéristiques hydrodynamiques et électrocinétiques de ces capillaires ont été étudiées, donnant une longueur de glissement de 23 µm et des efficacités de séparation électrophorétique 2 fois supérieures à celles obtenues sur un capillaire de silice vierge dans les mêmes conditions électrophorétiques. Le second chapitre étudie la génération de microbulles d’air dans des capillaires superhydrophobes. ). Les paramètres expérimentaux (tension, rayonnement UV, marqueur, revêtement superhydrophobe) nécessaires à l’obtention de ces microbulles ont été identifiés). Ces bulles ont été caractérisées (diamètre ~35-39 µm ; longueur ~10 mm ; potentiel zeta ~ -62.6 mV). Le troisième chapitre propose une méthodologie expérimentale, basée sur la théorie de l’électrochromatographie, permettant d’évaluer l’adsorption résiduelle de protéines sur la paroi des capillaires. Cette nouvelle approche présente deux intérêts : (i) pouvoir comparer les performances séparatives sur différents greffages via l’adsorption résiduelle, et (ii) optimiser les paramètres expérimentaux (longueur, diamètre interne, tension appliquée) afin de minimiser l’impact de l’adsorption sur les efficacités de séparationThe main objective of this thesis is to study different approaches for the modification of the electrophoresis capillary intern wall to enhance separation efficiency and reproducibility for biomolecules (model peptides and/or proteins) in acidic conditions. The first chapter (outside of bibliographic study) is dedicated to superhydrophobic coatings study. The goal is to prevent analytes adsorption by suppressing any interaction between the superhydrophobic wall and the analytes (anti-wettability). An original coating process has been developed to obtain superhydrophobic capillaries by studying the influence of layers number, coating nature, and filling and flushing pressure during layers deposition. Superhydrophobic coatings have been obtained, for the first time, with diameters from 50 µm to 180 µm. Hydrodynamic and electrokinetic characteristics have been studied, giving slipping length of 23 µm and efficiency separation increased twofold compared to fused silica capillary in the same electrophoretic conditions. The second chapter studies an air microbubbles generation process using superhydrophobic capillaries. The experimental parameters (voltage, UV ray, marker, superhydrophobic coating) needed to obtain those bubbles have been identified. Those bubbles have been characterized (diameter ~35-39 µm; length ~10 mm; zeta potential ~ -62.6 mV). The third chapter offer an experimental methodology, based on the electrochromatography theory, allowing to evaluate the residual adsorption of proteins on the capillary wall. This approach have two interesting points: (i) allowing to compare separative performances of different coatings via residual adsorption, and (ii) optimizing the experimental parameters (length, internal diameter, applied voltage) to minimize the impact of adsorption on the separation efficiencies
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Baratuci, William Brian. "Counteracting flow electrophoresis". Case Western Reserve University School of Graduate Studies / OhioLINK, 1991. http://rave.ohiolink.edu/etdc/view?acc_num=case1055352218.
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Ruel, Coralie. "Apport de l'électrophorèse capillaire pour l'étude des anomalies de glycosylation de protéines liées à des pathologies : vers l'identification de nouveaux biomarqueurs". Thesis, Université Paris-Saclay (ComUE), 2018. http://www.theses.fr/2018SACLS519.
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La glycosylation est l’une des principales modifications post-traductionnelles des protéines. La glycosylation des protéines est fortement modifiée lors de diverses pathologies comme le cancer, la polyarthrite rhumatoïde ou les troubles congénitaux de la glycosylation («Congenital disorders of glycosylation», CDGs). Ainsi, la nature et les proportions relatives des oligosaccharides liés aux protéines peuvent être utilisées pour dépister, pronostiquer voire suivre l’évolution de pathologies. Les travaux de cette thèse se sont focalisés sur l’étude de la glycosylation pour permettre le dépistage de certaines pathologies : les CDGs, la maladie d’Alzheimer (MA) et la dégénérescence rétinienne. Plusieurs stratégies fondées sur l’électrophorèse capillaire (EC) ont été envisagées pour répondre à cet objectif. Tout d’abord, le développement d’une méthode par EC de zone (ECZ) l’apolipoprotéine C-III (ApoC-III) intacte, une O-glycoprotéine impliquée dans le dépistage des troubles de la O-glycosylation, a permis de séparer les différentes glycoformes selon leur degré de sialylation. L’analyse d’échantillons d’ApoC-III standard provenant de différents fournisseurs par spectrométrie de masse (MS) MALDI -TOF a mis en évidence une hétérogénéité liée à la présence (inattendue) de formes carbamylées. Un traitement du plasma basé sur une immunocapture de l’ApoC-III suivie d’une dérivation sur billes magnétiques de la protéine par un fluorophore a permis de séparer ses différentes glycoformes en ECZ. Ensuite, une analyse N-glycomique de fluides biologiques employant une nouvelle technique de préparation d’échantillon que nous avons adaptée au plasma et au liquide céphalorachidien, a été réalisée par EC en gel couplée à une détection par fluorescence induite par laser (ECG-LIF) sur des patients sains et atteints de la MA. Cette étude a permis de mettre en évidence quelques modifications des N-glycanes chez ces patients. Enfin la combinaison des deux stratégies d’analyse de la glycosylation (glycoprotéine intacte et glycanes libérés), a permis de détecter la transferrine intacte présente dans le liquide vitré à des concentrations faibles ainsi que ses N-glycanes libérés, dans le cadre du dépistage d’une maladie oculaire. L’EC-QTOF-MS a également été explorée pour l’analyse de N-glycanes dérivés avec un nouveau fluorophore permettant d’améliorer la sensibilité en MSGlycosylation is one of the most main types of post-translational modifications of proteins. Disease-associated modifications in protein glycosylation have been observed for various pathologies such as cancer, rheumatoid arthritis, or «Congenital disorders of glycosylation» (CDGs). They are often exploited for diagnosis, prognosis and monitoring of these diseases. This thesis work focused on the glycosylation study with the aim to allow the screening of different pathologies: CDGs, Alzheimer’s disease and retinal degeneration diseases. Different strategies based on capillary electrophoresis (CE) were considered to fulfil this goal. First, the developement of a CZE analysis of intact apolipoprotein C-III (ApoC-III), an O-glycoprotein implied in the screening of O-glycosylation disorders, allowed the separation of its glycoforms according to their sialylation degree. The MALDI-TOF mass spectrometry (MS) analysis of standard ApoC-III batches from different standard ApoC-III batches from different suppliers highlighted an additonnal heterogeneity due to (unexpected) carbamylated species. A plasma pretreatment based on an immunocapture of apoC-III followed by protein derivatization on magnetic beads using a fluorophore allowed to separate its glycoforms by CZE. Second, a glycomic analysis of biologicial fluids using a new sample treatment method that we adjusted to plasma and cerebrospinal fluid samples was performed by CGE-LIF on controls and Alzheimer’s patients. It allowed to highlight some modifications of N-glycans for this disease. Finally, the combination of both strategies of glycosylation analysis (intact glycoprotein and released N-glycans) allowed the detection of intact transferrin present in vitrous humor but also of its released N-glycans for the screening of retinal degeneration disease. CE-QTOF-MS was also investigated for the analysis of released N-glycans derivatized by a new fluorophore which increases MS sensitivity
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Zha, Wuyi. "Velocity-difference induced focusing in capillary electrophoresis and preparative capillary electrophoresis". Thesis, University of British Columbia, 2007. http://hdl.handle.net/2429/448.
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Velocity-difference induced focusing (V-DIF) with a dynamic pH junction in capillary electrophoresis (CE) using a sample with a pH different from that of the background electrolyte (BGE) was developed in our group, but the mechanism was not well understood. In this work, the mechanism of this focusing technique was first studied using an appropriate dye to monitor the pH of the BGE and the sample during the focusing process. A mechanism was proposed based on the experimental results. This technique was then applied to serotonin to improve the detection limit when CE was used with a UV absorption detector. It was also applied to focus amino acids, peptides, and proteins to improve the concentration sensitivity. It is found that the pKa rather than the pI of the analytes is the key criterion for selecting the pH for the sample and for the BGE to obtain the optimum focusing for these molecules. Since UV detection only provides migration time information, more structure information is obtained by using a photodiode array (PDA) and mass spectrometer (MS) for peak identification. Comparisons were made between the PDA detection and MS detection for aromatic amino acids with V-DIF using a dynamic pH junction. This V-DIF technique was then applied to non-aromatic amino acids with MS detection. It was used at low pH with positive ESI-MS detection and at high pH with negative ESI-MS ionization. The results of the two methods were compared and discussed. Finally, the preparative operation of continuous flow counterbalanced CE (FCCE) was studied. The effects of larger sample volumes and multiple capillary systems on improving the purification yield were investigated.
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Xu, Aoshuang. "Development in electrophoresis instrumentation for two-dimensional gel electrophoresis of protein separation and application of capillary electrophoresis in micro-bioanalysis /". [Ames, Iowa : Iowa State University], 2008.
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Liang, Yufu. "Chiral Separation Using Capillary Electrophoresis (CE) and Continuous Free Flow Electrophoresis (CFFE)". University of Cincinnati / OhioLINK, 2003. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1067615432.
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McCormack, Kathleen Anne. "Capillary electrophoresis and electrochromatography". Thesis, University of Edinburgh, 1991. http://hdl.handle.net/1842/11108.
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Yurdakul, Saruhan. "Electrophoresis of electrogenerated bubbles". Thesis, Imperial College London, 1991. http://hdl.handle.net/10044/1/58542.
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The fundamental understanding of the interfacial charge on gas bubbles and the consequences of such charge are essential in understanding the behaviour of physicochemical systems involving liquid/gas and solid/liquid/gas interfaces. Such interfaces are involved in many industrial processes such as electrolytic gas evolution, particle flotation and bubble coalescence. The knowledge of such interfaces will aid mass transfer calculations. This thesis describes the application of a laser Doppler anemometer (LDA) system to the measurement of bubble electrophoretic mobilities, giving a measure of adsorbed charge. Single bubbles were electrogenerated in surfactant-free electrolytes, characterised by bubble rise rates, and their behaviour investigated in an electric field applied parallel to the direction of rise, so that, depending on the field direction, an increase or a decrease in the rise velocities was obtained. This field orientation served to decouple the hydrodynamic and field-induced charge polarisation. The velocity measurements using LDA showed a large degree of scatter despite numerous modifications to the optics and the signal processing. This culminated in the belief that a double LDA system was necessary to optimise the reliability and accuracy of the technique. Measurements using a Kodak high speed camera and recording system showed that the bubbles were negatively charged over the pH range studied (3-11), as indicated by their migration towards the anode under the influence of an applied electric field, with mobilities showing a radius and field dependence, implying that the adsorbed charge at the gas/electrolyte interface was mobile and polarisable. Large mobilities (10-60 x 10"® m2 s"^ V"^) were observed in comparison with results from previous bubble electrophoresis experiments with lateral fields. This was explained in terms of the enhanced charge polarisation occurring in the parallel electric field to the rise vector. A qualitative explanation for the decoupling of the hydrodynamic and field-induced charge polarisation has also been provided. In a separate series of experiments, under sufficient field conditions to overcome buoyancy forces, rising bubbles were stopped and held stationary. It was shown by extrapolation that bubbles possessed an iso electric point between pH 2 and 3, being positively charged below pH 2 and negatively charged above pH 3, supporting the hypothesis that the preferential adsorption of OH /H+ ions gives rise to the net charge. A laser reflection technique was investigated to measure the thickness of a liquid film formed between a bubble and the planar gas/electrolyte interface when they are in close proximity of each other. Preliminary investigations on macroscopic soap films showed the technique to be suitable for studying film thinning rates, though further refinement is necessary to study microscopic transient films. Electrophoresis measurements using a high speed camera have shown that bubbles preferentially adsorbed OH-/H+ ions from the solution in the absence of surfactants. This charge resided on a highly mobile interface and could be polarised by the actions of the hydrodynamics and the electric field. The laser Doppler anemometer system requires further development to achieve more accurate bubble velocity profiles in order to detect the small changes that occur.
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SPIESER, ESTELLE. "Electrophorese capillaire : applications pharmaceutiques". Strasbourg 1, 1994. http://www.theses.fr/1994STR15067.
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Pálmarsdóttir, Sveinbjörg. "Procedures for sample clean-up and concentration in capillary zone electrophoresis for determination of drugs in biosamples". Lund : Dept. of Aanalytical Chemistry, University of Lund, 1996. http://catalog.hathitrust.org/api/volumes/oclc/38045310.html.
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McKillop, Andrew G. "Ion mobilities in capillary electrophoresis". Thesis, Loughborough University, 1996. https://dspace.lboro.ac.uk/2134/28235.
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The project has investigated the electrophoretic mobilities of sets of model compounds to determine the effects of size and shape on ion mobilities. Methods were developed for the analysis of compounds in order to quote accurate electrophoretic mobilities. Using the obtained electrophoretic mobilities mobility orders were correlated with structural properties.
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Koeble, Konrad. "Electrophoresis of megabase DNA molecules". Thesis, University of Oxford, 1991. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.302890.
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Penn, Sharron Gaynor. "Chiral analysis by capillary electrophoresis". Thesis, University of York, 1994. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.241074.
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Sohn, Dosung. "Invertebrates analysis by capillary electrophoresis". [Gainesville, Fla.] : University of Florida, 2009. http://purl.fcla.edu/fcla/etd/UFE0024340.
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Caubert, Florent. "Etude de l'imprégnation électrophorétique, en milieu aqueux, de nanoparticules de boehmite, en vue du colmatage d'un film anodique poreux sur alliage d'aluminium 1050". Thesis, Toulouse 3, 2016. http://www.theses.fr/2016TOU30394/document.
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Les pièces en aluminium sont largement utilisées dans le domaine aéronautique en raison de leurs bonnes propriétés mécaniques. Mais elles nécessitent un traitement de surface pour améliorer leur tenue en corrosion. Soumises à de nouvelles normes sur l'utilisation de produits chimiques et à la prise de conscience de la protection environnementale et humaine, les industries aéronautiques doivent à présent impérativement remplacer les procédés de traitements de surface actuels, devenus obsolètes car incluant des composés CMR. L'objectif de ces travaux de recherche est de développer un traitement de surface par voie liquide, à la fois innovant et conforme à la législation REACH, pour améliorer les propriétés d'anticorrosion des alliages d'aluminium ; le procédé d'élaboration présentement étudié, est composé d'une anodisation poreuse puis d'un colmatage par imprégnation de particules au sein des pores. Un film anodique poreux " modèle " a tout d'abord été élaboré et caractérisé : son épaisseur est de 10 µm, tandis que les pores sont rectilignes et ont un diamètre moyen de 120 nm. Puis, nous avons étudié la synthèse par voie aqueuse, de nanoparticules de boehmite, l'optimisation des différents paramètres de synthèse ayant permis finalement d'obtenir des particules d'une taille inférieure à celle des pores du film anodique. Deux techniques d'incorporation ont ensuite été expérimentées : le trempage-retrait et l'électrophorèse. La compréhension des mécanismes mis en jeu et de l'influence de différents paramètres opératoires, a permis une maitrise des procédés et l'insertion effective de particules. Des caractérisations microstructurales ont en particulier montré que l'insertion de particules est plus aisée dans le cas d'une électrophorèse avec une tension pulsée. Enfin, la mise en œuvre d'un post-traitement hydrothermal après l'imprégnation, a permis d'obtenir un colmatage complet des pores du film anodique, et d'augmenter significativement les propriétés anticorrosionAluminum parts are widely used in the aeronautical field because of their good mechanical properties. But they require a surface treatment to improve their resistance to corrosion. Subject to new standards on the use of chemicals and awareness of environmental and human protection, the aeronautical industry must now replace current surface treatment processes, which have become obsolete because they include CMR compounds. The aim of this research is to develop a surface treatment, both innovative and REACH compliant, to improve the anticorrosion properties of aluminum alloys; the process here studied, is composed of a porous anodization and a sealing by impregnation of particles within the pores. A "model" porous anodic film was first prepared and characterized: its thickness is 10 µm, while the pores are straight and have a mean diameter of 120 nm. Then, we studied the aqueous synthesis of boehmite nanoparticles; the optimization of the synthesis parameters finally allowed to obtain a particle size smaller than the pore diameter. Two incorporation techniques were then tested: dip-coating and electrophoresis. The understanding of the involved mechanisms and of the influence of different operating parameters, allowed a control of the processes and the effective insertion of particles. In particular, microstructural characterizations showed that the particle insertion is easier using pulsed voltage electrophoresis. Finally, a hydrothermal post-treatment after the impregnation, allowed to obtain a complete sealing of the anodic film pores, and to significantly increase the anticorrosion properties
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McLeod, George Slater. "Development of capillary electrophoresis and capillary electrophoresis-mass spectroscopy methods for application in food analysis". Thesis, University of East Anglia, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.267331.
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Patricio, Magalhaes Candida Ana. "Développement d'un procédé couplé sol-gel/électrophorèse pour des applications en anti-corrosion". Thesis, Toulouse 3, 2016. http://www.theses.fr/2016TOU30343/document.
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L'objectif de ces travaux de thèse a été de développer un système de protection anticorrosion sur alliages d'aluminium 2024 en combinant le procédé sol-gel à la technique de dépôt par électrophorèse (ou EPD). Dans cette optique, nous nous sommes intéressés à l'évaluation du couplage de la technique de dépôt par EPD, technique offrant l'avantage de réaliser des dépôts à la fois épais, couvrants et homogènes en épaisseur même sur pièces complexes, au procédé sol-gel, bien connu et fortement prometteur pour développer des revêtements anti-corrosion performants. Le travail de thèse s'est focalisé sur la réalisation de dépôts par électrophorèse à partir d'un sol hydride, chargé ou non en particules de boehmite (NPs), et sur l'étude de différents paramètres électrophorétiques (paramètres à la fois liés au medium et au procédé lui-même). Nous avons ensuite cherché à connaître les propriétés des revêtements mis en forme par EPD (propriétés anti-corrosion et perméabilité des films en particulier) via des caractérisations électrochimiques (étude par Spectroscopie d'Impédance Electrochimique (SIE), électrode à disque tournant (EDT)). Ces dernières études ont été réalisées sur alliage d'aluminium 2024 (AA2024), dans un premier temps, puis sur substrat modèle (substrat inerte tel que l'or (Au)) dans le but d'évaluer la réponse électrochimique intrinsèque du revêtement. Dans un premier temps, nous avons pu obtenir par EPD et de façon répétable, des dépôts homogènes et d'épaisseurs contrôlées sur AA2024 et Au. La croissance de ces dépôts est à la fois le résultat de la migration des NPs et de l'augmentation du pH interfacial, provoqué par l'électrolyse de l'eau mais responsable, à terme, d'une redissolution du film. Ce phénomène de redissolution/décollement a pu néanmoins être limité en modulant la forme du signal électrique (application d'un potentiel périodique). Aussi, nous avons démontré que la quantité de NPs présente dans le sol précurseur joue un rôle clé dans les épaisseurs de dépôts obtenues (des épaisseurs dans la gamme 7 à 20 µm ont pu être atteintes en doublant uniquement la concentration en NPs vectrices dans le sol précurseur). En ce qui concerne le comportement électrochimique des différents systèmes mis en forme par EPD, la modélisation des spectres SIE, via le logiciel Z view, montre que les propriétés barrières des revêtements chargés tendent à se dégrader plus vite avec le temps d'immersion que pour ceux obtenus sans particules. Nous avons finalement cherché à mesurer la perméabilité des films électrophorétiques via un dispositif de type électrode à disque tournant (EDT). Les résultats obtenus ont montré que le coefficient de perméabilité kf des films électrophorétiques évolue fortement, dès les premières heures d'immersion, et augmente d'autant plus que le système est chargé en NPsThe realisation of organic/inorganic coatings on metal substrates, prepared by sol-gel route and shaped by electrophoretic deposition (EPD), is a new combined process which has been the subject of only few studies. EPD technique offers an easy control of the thickness and morphology of the film even on substrates of complex shape, which are the main challenges for all kinds of deposition techniques used in various industrial fields. Moreover, sol-gel route has been extensively studied as a potential alternative pre-treatment to prepare a variety of materials with versatile applications from anti-corrosion to anti-wear. So, coupling these two techniques is one way to obtain both benefits on a same system. In this work, the electrophoretic deposition was performed on AA2024 from an aqueous sol suspension containing sol-gel boehmite nanoparticles (NPs). The influence of the applied voltage and deposition time on the deposit thickness was studied. The effect of the concentration of NPs, added in the precursor sol, on the thickness was also investigated. It is shown that an increase in the applied voltage and deposition time increased the thickness of the deposit film (from 2 to 11 µm). However, for a same voltage, increasing the concentration of NPs in the precursor sol, progressively increases the coating thickness (Figure 1.) and appears as a key parameter to adjust the coating thickness. Finally, it was demonstrated that a perfect control of the microstructure and the deposit thickness was achievable, thanks to both EPD parameters and sol properties. The electrochemical properties of the electrophoretic coatings are then studied by Electrochemical Impedance Spectroscopy (EIS) and by rotating disk electrode and showed that the permeability of the coatings increased with the particles concentration
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Hütter, Gero. "Chemoresistenzassoziierte Veränderungen der Proteinexpression bei Kolon-, Mamma-, Magen-, Pankreaskarzinom und Fibrosarkom mit Hilfe der hochauflösenden zweidimensionalen Elektrophorese im immobilisierten pH-Gradienten". Doctoral thesis, Humboldt-Universität zu Berlin, Medizinische Fakultät - Universitätsklinikum Charité, 2000. http://dx.doi.org/10.18452/14467.
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Die Therapie von disseminierten malignen Tumoren durch Chemotherapie allein oder in Kombination mit Bestrahlung oder Hyperthermie führt nur in 20-50% zu einem Ansprechen. Dieser Behandlungserfolg wird häufig dadurch limitiert, daß im Verlauf der Therapie zur Entwicklung einer Chemoresistenz kommt. Ziel der Arbeit war es, eine generelle Analyse der Proteinexpression in der in vitro herbeigeführten Chemoresistenz durchzuführen. Dafür wurden Zellkulturen von Magen-, Kolon-, Pankreas, Mammakarzinom und Fibrosarkom die eine Resistenz gegen Daunorubicin und Mitoxantron besitzen benutzt und mit der zweidimensionalen Gelelektrophorese im immobilisierten pH-Gradienten (pH 4,0-8,0) in der ersten Dimension und einen linearen Polyarcylamidgel (12%) in der zweiten Dimension analysiert. Nach Färbung in Coomassie blau wurde eine rechnergestützte Imageanalyse mit dem PDQuest System durchgeführt. Proteinspots die eine auffällige Änderung zeigten wurden isoliert, und nach enzymatischer Gelhydrolyse mikrosequenziert bzw. durch Massenspektrometrie und anschließender micropore HPLC Analyse identifiziert. Acht Proteine, die in den chemoresistenten Zellinien überexprimiert waren, konnten identifiziert werden: Thioredoxin, Annexin 1; Cofilin, Stratifin(14-3-3sigma), Rho-GDP-Dissoziations Inhibitor, Fettsäurebindendes Protein(E-FABP), Adenin Phosphoribosyl Transferase, und BCSG-1The therapy of advanced cancer using chemotherapy alone or in combination with radiation or hyperthermia yields an overall response rate of about 20-50%. This success is often marred by the development of resistance to cytostatic drugs. The aim was to study the global analysis of protein expression in the development of chemoresistance in vitro. We therefore used a cell culture model derived from the gastric, colorectal, pancreatic, mamma carcinoma and fibrosarcoma cell line selected to daunorubicin and mitoxantrone. These cell lines were analysed using two-dimensional electrophoresis in immobilized pH-gradients (pH 4.0-8.0) in the first dimension and linear polyacrylamide gels (12%) in the second dimension. After staining with coomassie brilliant blue, image analysis was performed using the PDQuest system. Spots of interest were isolated using preparative two-dimensional electrophoresis and subjected to microsequencing after enzymatic hydrolysis in gel, mass spectrometric data and sequencing of the peptides after their fractionation using microbore HPLC identified. Eight proteins were identified that were overexpressed in chemoresistant cell lines: Thioredoxin, Annexin 1, Cofilin, Stratifin(14-3-3sigma), Rho-GDP-dissoziation Inhibitor, fatty acid binding protein(E-FABP), adenin phosphoribosyl Transferase, and BCSG-1.
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Yu, DONGHUI. "Development of magnetic particle based biosensors and microreactors for drug analysis and biotransformation studies". Doctoral thesis, Universite Libre de Bruxelles, 2008. http://hdl.handle.net/2013/ULB-DIPOT:oai:dipot.ulb.ac.be:2013/210517.
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In the first part of this work, magnetized nanoporous silica based microparticles (MMPs) are used for horseradish peroxidase (HRP) immobilization and applied in amperometric peroxidase-based biosensors. A homemade magnetized carbon paste electrode permits the MMPs attraction close to the electrode surface. The resulting original biosensor is applied to the investigation of enzymatic oxidation of model drug compounds namely, clozapine (CLZ) and acetaminophen (APAP) by HRP in the presence of hydrogen peroxide. The biosensor operates at a low applied potential and the signal corresponds to the electro-reduction of electroactive species enzymatically generated. The biosensor allows performing the quantitation of the two drug compounds in the micromolar concentration range. It allows also the study of thiol compounds based on the inhibition of the biosensor response. Interestingly, distinct inhibition results are observed for HRP entrapped in the silica microparticles compared to the soluble HRP.We expect that this type of biosensors holds high promise in quantitative analysis and in biotransformation studies of drug compounds.
In the second part of this thesis work, HRP immobilized magnetic nanoparticles are injected on-line and magnetically retained, as a microreactor, in the capillary of a CE setup. The purpose of such a configuration is to develop an analytical tool for studying “in vitro” drug biotransformation. The advantages expected are (i) minimum sample (drug compound) and biocomponent (enzyme) consumption, (ii) high analysis throughput, (iii) selectivity and sensitivity. In order to illustrate the potential of such an instrumental configuration, it has been applied to study acetaminophen as model drug compound. The mechanistic information obtained by the HRP/H2O2 system is in agreement with literature data on acetaminophen metabolization. Horseradish peroxidase kinetic studies are realized by this setup and the apparent Michaelis constant is determined. Capillary electrophoresis permitted the identification of APAP off-line biotransformed products such as N-acetyl-p-benzoquinone imine (NAPQI), the APAP dimer and APAP polymers as inferred from literature data. The formation of the APAP dimer was further confirmed by electrospray ionization mass spectrometry.
Doctorat en Sciences biomédicales et pharmaceutiques
info:eu-repo/semantics/nonPublished
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Richards, Dawn P. "Electrophoretic separations of biopolymers". Thesis, National Library of Canada = Bibliothèque nationale du Canada, 2000. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape3/PQDD_0013/NQ59661.pdf.
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Branco, Marta Sofia de Pinho. "Electrophoretic deposition of kaolin". Master's thesis, Universidade de Aveiro, 2017. http://hdl.handle.net/10773/22591.
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Mestrado em Engenharia de MateriaisA deposição eletroforética (EPD) é uma técnica interessante do ponto de vista de processamento de materiais permitindo a formação de filmes dos mais variados materiais em diferentes substratos condutores. É uma técnica simples, versátil e de baixo custo associado. No presente trabalho, conduziram-se estudos que permitiram estabelecer as condições apropriadas para a produção de filmes espessos de caulino por EPD. Depositaram-se filmes de caulino em verde, com cerca de 5 mg, em três tipos de substrato: aço inoxidável, folha de platina e silício platinizado. Para a identificação das condições de deposição, contribuiu o estudo sistemático das condições de preparação da suspensão de caulino que incluíram a avaliação do potencial zeta de suspensões com o pH da mesma, do efeito da composição do meio suspensor (água, etanol, ou mistura de ambos), do recurso ao iodo como aditivo, e da variação com o tempo da transmitância da luz UV por parte dos diferentes meios suspensores, com ou sem iodo. Os resultados obtidos permitiram concluir que as melhores condições de deposição são as que combinam a adição de alguma água ao etanol, enquanto meio suspensor, e o uso de iodo como aditivo. Os filmes preparados por EPD foram sinterizados a 1200 e 1300 oC, durante 2 h. A microestrutura dos filmes, antes e após sinterização, observada por microscopia eletrónica de varrimento (SEM) permitiu concluir que as partículas de caulino tendem a depositar de uma forma orientada em que as suas superfícies basais se alinham paralelamente ao substrato. Os filmes de caulino sinterizados foram submetidos a ensaios de nanoindentação e determinou-se a sua dureza Vickers e módulo de Young para os quais se obtiveram, respetivamente, 300 MPa e 40 GPa. Este trabalho contribuiu para identificar condições para obter filmes espessos de caulino de cuja a microestrutura anisotrópica se aponta a possibilidade de aceder a propriedades maximizadas segundo determinadas direções o que do ponto de vista das suas aplicações pode abrir novas oportunidades.
Electrophoretic deposition (EPD) is an interesting technique from the point of view of materials processing. The technique allows the formation of films of many different materials on different conductive substrates. Besides that, EPD is a simple, versatile and low cost technique. The studies conducted, in the present work, allowed the establishment of appropriate conditions to produce kaolin thick films by EPD. Green kaolin films with around 5 mg were deposited on three types of substrate: stainless steel, platinum foil and platinized silicon. A systematic study about the preparation conditions of the kaolin suspension contributed to identify the deposition conditions. This study included the assessment of the pH dependence of zeta potential of the suspension and the effect of the suspension media (water, ethanol or a mixture of both) as well as the use of iodine as additive. Transmittance variation of the UV light with time was also assessed for the different suspension media with and without iodine. The obtained results allowed to conclude that the best deposition conditions are those that combine the use some water in the ethanol based suspension media added also with iodine. The kaolin films produced by EPD were sintered at 1200 and 1300 oC for 2 h. The observation of the films microstructure by scanning electron microscopy (SEM), before and after sintering, allowed to conclude that the kaolin particles tend to deposit in an oriented way in which their basal surfaces align parallel to the substrate. The sintered kaolin films were submitted to nanoindentation tests and their Vickers hardness and Young’s modulus was determined as 300 MPa and 40 GPa, respectively. This work contributed to identify the conditions to obtain kaolin thick films of which the anisotropic microstructure is expected the possibility of assessing maximized properties under certain directions. From the point of view of applications, this can open new possibilities
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Bohlin, Maria E. "Capillary electrophoresis of β2-glycoprotein I". Licentiate thesis, Karlstads universitet, Fakulteten för teknik- och naturvetenskap, 2006. http://urn.kb.se/resolve?urn=urn:nbn:se:kau:diva-3826.
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Mironov, Gleb. "Capillary Electrophoresis - Mass Spectrometry for Bioanalysis". Thesis, Université d'Ottawa / University of Ottawa, 2015. http://hdl.handle.net/10393/33004.
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Bioanalysis is a subdivision of analytical chemistry and deals with biological analytes such as metabolites, proteins, nucleic acids, small molecules, virus particles and entire cells. The rationale of my thesis was to achieve two goals: (i) develop a set of ready to use methods (ii) which are capable providing exact concentrations of analytes as well as kinetic and thermodynamic parameters of their interactions.
To investigate interactions between biomolecules special conditions are required which do not interfere with the course if biomolecule interactions. Establishing these conditions and optimization of separation and detection parameters can be tedious and can take longer than actual analysis of samples. I developed a variety of Capillary Electrophoresis – Mass Spectrometry (CE-MS) methods suitable for bionalalysis.
CE-MS establishes a new paradigm that separation methods together with MS detection can be used as comprehensive kinetic tools. Most previous attempts to use chromatography and electrophoresis for studying nucleic acid interactions were restricted to assuming slow or no equilibrium between reactants. Kinetic CE (KCE) shows that non-zero kinetics and structural dynamics must be taken into account when separation happens. KCE-MS could be a valuable supplement to IM-MS due to the separation of ions in solution according to their size-to-charge ratio.
These methods allowed to reveal new facts about biomolecules and added novel data to the bank of the mankind knowledge. For the best of my knowledge, kinetic parameters for TG2 and thrombin G-quadruplex folding were reported for the first time. I developed a homogeneous method to determine kon, koff and Kd of fast and weak noncovalent interactions between multiple unlabeled ligands (small molecule drugs) and an oligosaccharide (α- or β-cyclodextrin) simultaneously in one capillary microreactor. It has been shown for the first time that KCE can be used to separate and detect the slowly interconverting open and closed conformations of human TG2. It allowed the first direct measurement of the Kd value for calcium binding. Sixteen new substrates were discovered for three aminotransferases (AAT, BCAT, and DAAT). In addition, Viral qCE showed a feasibility to analyse both the count of intact viral particles and sample nucleic acid contamination.
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Salimi-Moosavi, Mir Mohammad Hossein. "Capillary electrophoresis in pure nonaqueous solvents". Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1997. http://www.collectionscanada.ca/obj/s4/f2/dsk3/ftp04/nq24045.pdf.
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Zhang, Zheru. "Single cell analysis by capillary electrophoresis". Thesis, National Library of Canada = Bibliothèque nationale du Canada, 2000. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape2/PQDD_0012/NQ59703.pdf.
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Cui, Huanchun. "Nonlinear electrophoresis in networked microfluidic chips". Online access for everyone, 2007. http://www.dissertations.wsu.edu/Dissertations/Fall2007/h_cui_110207.pdf.
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Xin, Yao. "Electrokinetic Modeling of Free Solution Electrophoresis". Digital Archive @ GSU, 2007. http://digitalarchive.gsu.edu/chemistry_diss/18.
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Modeling electrophoresis of peptides, proteins, DNA, blood cells and colloids is based on classical electrokinetic theory. The coupled field equations-Poisson, Navier-Stokes or Brinkman, and ion transport equations are solved numerically to calculate the electrophoretic mobilities. First, free solution electrophoretic mobility expressions are derived for weakly charged rigid bead arrays. Variables include the number of beads (N), their size (radius), charge, distribution (configuration), salt type, and salt concentration. We apply these mobility expressions to rings, rigid rods, and wormlike chain models and then apply the approach to the electrophoretic mobilities and translational diffusion constants of weakly charged peptides. It is shown that our bead model can predict the electrophoretic mobilities accurately. In order to make the method applicable at higher salt concentrations and/or to models consisting of larger sized subunits, account is taken of the finite size of the beads making up the model structure. For highly charged particles, it is also necessary to account for ion relaxation. This ion relaxation effect is accounted for by correcting "unrelaxed" mobilities on the basis of model size and average electrostatic surface, or "zeta" potential. With these corrections our model can be applied to the system with absolute electrophoretic mobilities exceeding approximately 0.20 cm2/kV sec and also models involving larger subunits. This includes bead models of duplex DNA. Along somewhat different lines, we have investigated the electrophoresis of colloidal particles with an inner hard core and an outer diffusive layer ("hairy" particles). An electrokinetic gel layer model of a spherical, highly charged colloid particle developed previously, is extended in several ways. The charge of the particle is assumed to arise from the deprotonation of acidic groups that are uniformly distributed over a portion (or all) of the gel layer. Free energy considerations coupled with Poisson-Boltzmann theory is used to calculate the change of the local pKa of the acidic groups depending on the local electrostatic environment. Based on the modeling of electrophoresis and viscosity, we predict that the thickness of the gel layer decreases as the salt concentration increases. And only the outermost portion of the gel layer is charged.
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Mangin, Catherine M. "Analysis of carrageenans using capillary electrophoresis". Thesis, University of York, 2000. http://etheses.whiterose.ac.uk/14043/.
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This thesis reports the use of capillary electrophoresis (CE) for the analysis of carrageenans, anionic polysaccharides extracted from red seaweeds and widely used in the food industry for their gelling and thickening properties. The three main types, kappa, iota and lambda, differ in the number of sulfate groups and the presence or absence of a 3,6-anhydro bridge in the disaccharide residue repeat unit. CE separates analytes according to their charge to frictional coefficient ratios, therefore it is suitable to separate these biopolymers. In order to detect polysaccharides in CE, our approach consisted in derivatising the reducing ends of the saccharides by reductive arnination with a fluorophore, l-arninopyrene-3,6,8- trisulfonate (APTS). This allowed sensitive detection by laser induced fluorescence. Method development gave optimal conditions for separation using a polyvinyl alcohol coated capillary and a 25 mM ammonium acetate, pH 8.0 background electrolyte. The effects of changes of both instrumental parameters (temperature, injection mode, field strength) and, the composition of the BGE (concentration and pH) are reported, and explained in terms of the physical chemistry of the BGE and the biopolymers. The conditions of the derivatisation reaction were studied in order to minimise degradation due in particular to acid catalysis and to reduction of the reacting sites occurring in competition with derivatisation. Characterisation of the derivatised carrageenans by SEC-MALLS- RI was performed and showed that the extent of degradation occurring during the labelling reaction was a maximum of 40 % for kappa and 20 % for iota and lambda. The presence of the label APTS in excess and its reaction with the reagents during the labelling reaction produces peaks interfering with those from the carrageenan. A sample clean-up was therefore required before injection onto CEo A comparison was made of a range of clean-up procedures (centrifugation, dialysis, preparative SEC) to remove side products of the reaction and salts and to concentrate the carrageenans. Various seaweed extracts were analysed, including standards of carrageenans not available commercially. This study revealed that carrageenans are complex structures, and often occurring as hybrids between sUb-types. CE has the ability to characterise these hybrids, unlike spectroscopic methods which detect individual residues. When using actual food products, preliminary steps such as defatting and dialysis were found to be necessary to allow satisfactory detection of carrageenans. Finally the strategy for sample purification, derivatisation, clean-up and separation was successfully applied to additive mixtures used as raw materials in the food industry and to finished products (jelly, dairy products). CE has proved to be a fast and sensitive method to identify and provide semi-quantitative information on carrageenans present in such mixtures.
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BOUYER, FREDERIC. "Elaboration de materiaux ceramiques par electrophorese". Besançon, 1999. http://www.theses.fr/1999BESA2013.
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L'objectif de ce travail est d'examiner le potentiel de l'electrophorese pour realiser des structures ceramiques multicouches (sic/sic poreux) a proprietes mecaniques superieures aux monolithes correspondants (sic). L'etude s'est portee principalement sur la fabrication de materiaux en carbure de silicium (sic) en milieu organique. Pour fabriquer des depots compacts, l'etat de dispersion des suspensions doit etre optimise et aucun effet de sedimentation ne doit etre observe. L'utilisation de polymere adsorbe sur le sic (polyvinylpirrolidone (pvp)) permet d'atteindre ces objectifs de meme que la combinaison d'un sel multivalent (alcl#3) avec un polymere non adsorbe (polyvinylbutyral (pvb)). Au contact de l'electrode, nous avons observe dans le systeme alcl#3/pvb la formation d'un liant inorganique (al(oh)#3) qui assure la cohesion du depot et l'adhesion des particules sur les particules predeposees. Les reactions electrochimiques permettent egalement de reduire la charge electrique des particules afin d'eviter toute polarisation de l'electrode. Pour densifier ou rendre poreux des materiaux en sic apres frittage, des additifs doivent etre incorpores dans les formulations (respectivement b#4c + noir de carbone ou latex / amidon). Si la charge des particules secondaires est de meme signe que celle du sic, un effet d'entrainement est observe avec formation d'un depot collectif ou le taux d'incorporation des particules minoritaires est identique a celui dans la suspension de depart. Dans le cas contraire, l'heterocoagulation de la suspension destabilise la suspension et necessite une optimisation de la teneur en additif de charge. La resistance mecanique des materiaux monolithiques a ete sensiblement amelioree par l'incorporation de couches poreuses. Avec les particules de latex comme precurseur de porosite, la resistance a la rupture est de 330 mpa (220 mpa pour le monolithe). Les particules d'amidon se decomposent partiellement et n'ont pas permis d'atteindre des valeurs aussi elevees.
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Khaled, Maha Yehia. "Selectivity and detection in capillary electrophoresis". Diss., This resource online, 1994. http://scholar.lib.vt.edu/theses/available/etd-06062008-164943/.
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Zhang, Junge Foley Joe Preston. "Quantitative biopharmaceutical applications of capillary electrophoresis /". Philadelphia, Pa. : Drexel University, 2009. http://hdl.handle.net/1860/3166.
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Ric, Audrey Marie Amélie. "Caractérisation d'aptamères par électrophorèse capillaire couplée au séquençage haut-débit Illumina". Thesis, Toulouse 3, 2017. http://www.theses.fr/2017TOU30388/document.
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Les aptamères sont des oligomères d'ADN ou d'ARN simple brin qui, en se repliant sous forme de structures tridimensionnelles peuvent avoir des interactions fortes et spécifiques envers un certain nombre de cibles. L'objectif de cette thèse a été de compléter les études existantes sur l'utilisation de l'électrophorèse capillaire (CE) et les aptamères afin de mettre au point une méthode de sélection d'aptamères par CE couplée à la fluorescence induite par laser et le séquençage haut-débit Illumina. Dans un premier temps, nous avons mis au point une méthode de détection et de séparation par électrophorèse capillaire couplée à la double détection UV-LEDIF d'une banque d'ADN en interaction avec une cible : la thrombine. C'est un modèle déjà étudié pour lequel deux aptamères ont fait l'objet de publications. Nous avons utilisé l'aptamère T29 dans le cadre de notre étude car c'est celui qui présente la meilleure affinité. L'électrophorèse capillaire est un puissant outil analytique qui facilite l'efficacité de sélection des aptamères et précise la détermination des paramètres d'interactions. Nous avons ainsi pu déterminer la constante d'affinité KD par CE-UV-LEDIF sur le modèle de base : la thrombine. Par ailleurs, nous montrons également comment l'utilisation du tampon Tris peut dégrader un ADN simple brin en électrophorèse capillaire et nous proposons comme alternative l'utilisation d'un tampon sodium phosphate dibasique qui évite ce phénomène de dégradation. Enfin, nous expliquons la difficulté d'amplification par qPCR et PCR d'un aptamère comme le T29 ayant une structure en G-quadruplex. Nous avons montré que le séquençage haut-débit Illumina nous a permis de trouver une corrélation entre le nombre de molécules séquencées et le nombre de séquences obtenues. L'analyse des séquences obtenues montre une quantité importante (20%) de séquences de T29 qui ne correspondent pas à la séquence de cet aptamère. Cela prouve que les étapes de PCR et de séquençage haut débit pour la détection de G-quadruplex peuvent induire un biais dans l'identification de ces moléculesAptamers are oligomers of small single-stranded DNA or RNA which can have strong and specific interactions with some targets when they fold into three-dimensional structures. The objective of this thesis was to complete existing studies on the use of capillary electrophoresis in order to develop a method for the selection of aptamers by CE coupled to laser induced fluorescence and Illumina high-throughput sequencing. In a first step, we developed a method of detection and separation by capillary electrophoresis coupled with the double detection UV-LEDIF of a DNA library interacting with a target: thrombin. It is a model already studied and for which two aptamers have been published. We used aptamer T29 as part of our study because it has the best affinity. Capillary Electrophoresis is a powerful analytical tool that facilitates the selection efficiency of aptamers and specifies the determination of the interaction parameters. We thus were able to determine the affinity constant KD by CE-UV-LEDIF on the basic model: thrombin. Moreover, we also show how the use of Tris buffer can degrade single-stranded DNA during capillary electrophoresis and we propose as an alternative the use of a dibasic sodium phosphate buffer which avoids the phenomenon of degradation. Finally, we explain the difficulty of amplification by qPCR and PCR of an aptamer such as T29 with a G-quadruplex structure. We showed that the Illumina high-throughput sequencing allowed us to find a correlation between the number of sequenced molecules and the number of sequences obtained. Analysis of the sequences obtained shows a significant amount (20%) of T29 sequences which do not correspond to the sequence of this aptamer. This shows that the PCR and high-throughput sequencing steps for the detection of G-quadruplex can induce bias in the identification of these molecules
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Gratia, Severine. "La cardiotoxicité de la doxorubicine : une étude transcriptomique, protéomique et phosphoprotéomique". Thesis, Grenoble, 2011. http://www.theses.fr/2011GRENV032/document.
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La doxorubicine (DXR) est l’un des médicaments les plus efficaces en chimiothérapie, mais sonapplication clinique est limitée par ses effets cardiotoxiques. Malgré des décennies de recherche, sesmécanismes pathogéniques ne sont toujours pas entièrement compris. Il s’ensuit qu’aucun traitementsatisfaisant, curatif ou préventif, n’existe. Dans cette étude, nous recherchons les mécanismes designalisation cellulaire impliqués. Deux modèles expérimentaux de toxicités, aigue d’une part (coeurisolé et perfusé de rat avec la DXR), et chronique d’autre part (rat traité à la DXR), ont permis deréaliser une étude ciblée (sur les voies de signalisation énergétiques) et deux études systémiques(phosphoprotéomique et transcriptomique). Les résultats combinés de ces travaux ont montré que laDXR modifiait le niveau de phosphorylation (activation) ou l’expression génique de protéinesimpliquées dans trois domaines fonctionnels distincts : métabolisme énergétique, réponses au stress,et structure/fonction du sarcomère. (i) Métabolisme énergétique : nous avons confirmé la surprenanteinhibition de l’AMPK, probablement provoquée par un contrôle négatif exercé par des partenaires designalisation (Akt et ERK), plutôt que par une modification des kinases activatrices en amont. Nousavons également montré l’augmentation du niveau de phosphorylation de la PDH, ce qui, en inhibantl’enzyme, ralentit le cycle de Krebs. Cependant, nous avons également observé un phénomènecompensatoire de surexpression de gènes codant pour des enzymes de la glycolyse et du cycle deKrebs ; (ii) Réponses au stress : dans nos modèles, la DXR génère des stress énergétique,génotoxique et oxydatif. Cependant, seuls quelques mécanismes compensatoires sont activés (lesvoies de signalisation de DNA-PK–Akt–GSK3, diverses chaperonnes). Les autres semblent êtreinhibées suggérant que l’amoindricement des réponses au stress serait un des mécanismes de lacardiotoxicité de la DXR; (iii) Structure/fonction du sarcomère: L’augmentation de la phosphorylationde la desmine ainsi que la réduction du nombre de transcrits codant pour des protéines essentiellesau développement cardiaque normal pourraient être la cause de la désorganisation du réseaumyofibrillaire. En conclusion, ces résultats révèlent potentiellement de nouveaux mécanismes de lacardiotoxicité induite par la DXR et permettent d’envisager de nouvelles cibles moléculaires pour ledéveloppement de stratégies protectricesDoxorubicin (DXR) is an efficient anticancer drug, the use of which is limited by seriouscardiotoxicity. Despite decades of research, its pathogenic mechanisms are not fully understood, andefficient preventive or curative strategies are not available. Here we address the question whethermechanisms in cardiac cell signaling contribute to the toxicity phenotype. Using experimental modelsfor acute (DXR-perfused, isolated rat hearts) or chronic toxicity (rats injected with DXR), we conducteda targeted study (focusing on energy signaling pathways) and two non-biased studies(phosphoproteomics and transcriptomics). The combined data reveal DXR-induced alterations inphosphorylation (activation) status or gene expression of proteins in mainly three functional domains:energy metabolism, stress responses, and sarcomere structure. (i) Energy metabolism: We confirm aparadox inhibition of AMPK signaling, that is rather due to inhibitory cross-talk with related signalingpartners (Akt, ERK) than impaired AMPK upstream signaling. We also show, among others, theincrease of inhibitory phosphorylation of pyruvate dehydrogenase, slowing down Krebs cycle, but alsoa compensating upregulation of glycolysis and Krebs cycle enzyme transcripts. (ii) Stress-responses:In our models, DXR generates energetic, oxidative and genotoxic stress, but only some compensatorystress responses are activated (DNA-PK–Akt–GSK3 pathway, chaperones). Many others seem to beinhibited, suggesting a blunted response to stress as component of DXR toxicity. (iii) Sarcomerestructure/function: We detect increased phosphorylation of desmin and reduced transcripts essentialfor e.g. normal heart development as potential causes for a disorganized myofibrillar network. Inconclusion, these results reveal some novel potential mechanisms of DXR-induced cardiotoxicity andsuggest new targets for protective strategies
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Ngom, Sokhna Mery. "Dispositifs nanofluidiques à électro-préconcentration sélective". Thesis, Université Paris-Saclay (ComUE), 2019. http://www.theses.fr/2019SACLS459.
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Détecter des biomolécules à l’état de traces reste l’un des enjeux actuels des biopuces. Les dispositifs nanofluidiques apparaissent aujourd’hui comme une voie prometteuse pour simultanément concentrer et détecter des biomolécules. Cette électro-préconcentration est possible grâce au caractère de perméabilité sélective de la nanofente (ou du nanocanal), qui se comporte sous champ électrique comme un «super-filtre» moléculaire à perméabilité sélective. Ce nanofiltre permet de piéger les analytes en amont ou en aval de la fente, dans l’un ou l’autre des réservoirs (anodique ou cathodique). Au cours de ce doctorat, j’ai développé et étudié des dispositifs nanofluidiques sur la base de deux géométries différentes : des nanofentes horizontales uniques et des réseaux de nanocanaux verticaux, dans une géométrie de code-barres. Pour les nanofentes horizontales, j’ai étudié l’évolution de la conductance en fonction de la force ionique et de la géométrie de la nanofente. Sur la base d’un protocole d’électro-préconcentration assistée en pression, j’ai établi des diagrammes « champ électrique/pression » qui permettent de prédire l’obtention d’un point focal stable où les analytes vont se concentrer. J’ai étudié le rôle de la longueur de la nanofente sur l’observation de ce point focal pour deux molécules modèles, la fluorescéine et l’ovalbumine. Pour les dispositifs à code-barres, j’ai mis au point un procédé de nanostructuration par lithographie électronique couplée à de la gravure profonde et un protocole de collage verre-verre. Les profils d’électropreconcentration obtenus pour différentes nanofentes au sein des codes-barres dynamiques permettent de discuter du rôle de la géométrie sur l’observation du point focalDetecting trace biomolecules remains one of the current challenges for biochips. Nanofluidic devices appear today as a promising way to simultaneously concentrate and detect biomolecules. This electropreconcentration is possible thanks to the selective permeability of the fluidic nanoslit, which behaves under electric field as a molecular selective "super-filter". This nanofilter makes it possible to trap the analytes upstream or downstream of the slot, in one or the other of the reservoirs (anodic or cathodic). During this Ph.D., I developed and studied nanofluidic devices based on two different geometries: single horizontal nanoslits and vertical nanochannel arrays, in a barcode geometry. For horizontal nanoslits, I studied the evolution of the conductance as a function of the ionic strength and the nanoslit geometry. Based on a pressure-assisted electro-preconcentration protocol, I have established "electric field/ pressure" diagrams allowing predicting stabilization of a focal point where the analytes will concentrate. I have studied the role of the nanoslit length for two model molecules, fluorescein and ovalbumin. For barcode devices, I developed both a nanostructuration process by electron beam nanolithography coupled with deep etching and a glass-glass bonding protocol. The electroconcentration profils obtained for different nanofentes is discussed based on different dynamic barcodes
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Bowser, Michael T. "Dynamic complexation of solutes in capillary electrophoresis". Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1998. http://www.collectionscanada.ca/obj/s4/f2/dsk2/ftp02/NQ34516.pdf.
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Gerhardt, Geoff C. "Square-wave voltammetry detection for capillary electrophoresis". Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1999. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape7/PQDD_0015/NQ43512.pdf.
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Shim, Jaesool. "Modeling and simulation of non linear electrophoresis". Online access for everyone, 2007. http://www.dissertations.wsu.edu/Dissertations/Fall2007/j_shim_121407.pdf.
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Schrader, Jeffrey A. "A doppler electrophoresis instrument for macromolecular characterizations". Thesis, This resource online, 1990. http://scholar.lib.vt.edu/theses/available/etd-05022009-040443/.
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Hosseini, Seyed Homayoun. "Temperature gradient gel electrophoresis development and application". Diss., Georgia Institute of Technology, 1994. http://hdl.handle.net/1853/25614.
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Huang, Xingye. "Chiral separation of pharmaceuticals by capillary electrophoresis". Thesis, University of Nottingham, 2010. http://eprints.nottingham.ac.uk/11645/.
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Conventional capillary zone electrophoresis (CZE) methods, with simple buffer solute, natural or derivatized cyclodextrin and organic additive in BGE, have been developed for a group of ten standard chiral pharmaceutical compounds representing different physiochemical properties and pharmaceutical activities. In this study, factors affecting chiral separation in CZE, including BGE pH value, ionic strength, chiral selector type, selector concentration and organic additives, were optimized. A maximum of eight standard compounds were separated by three different standard methods which were developed. The electrophoretic behaviours of the standard compounds observed were in good agreement with the literature. Partial filling technique (PFT) was studied as a complementary approach to conventional CZE methods for enantioseparation of standard compounds. Partial filling time, selector type and concentration were investigated; a maximum of seven standard compounds were separated by optimized filling time and three different chiral selectors. However, for five of the separated pharmaceuticals, the chiral resolutions achieved were much lower than those obtained from conventional CZE methods. Key observations from the experiment were supported by previous research. For the first time, glycidol was evaluated as a covalently bonded coating material on CE capillary for enantioseparation. Hyperbranched polyglycidol brushes were grown directly from Si/SiO2 surface via anionic ring-opening polymerization, using surface Si-OH groups as initiator. This grafting-form technique eliminated the need for initiator functionalized self-assembled monolayers on the surface, and the thickness and complexity of the hyperbranched polymer brushes could be well controlled in this process. Polyglycidol coating was established on the surface of glass slides and then adapted to CE capillary. Both fused silica capillary and etched capillary were used to examine the electrophoretic properties of polyglycidol coating. Chiral polyglycidol coating was compared with the standard CZE method developed and showed excellent chiral selectivity for standard compounds. Nine out of ten standard compounds were separated with poly-S-glycidol coated capillary, using simple buffer solute containing organic additive. Application of etched capillary further improved the enantioseparation resolution and peak efficiency for those standard compounds. Stability and coating regeneration ability were studied. Polyglycidol coating developed on CE capillary gradually lost its chiral selectivity after 50 30-min runs with acidic BGE. Coating regeneration on the remaining surface was difficult. The result indicated that glycidol isomer can be used as monomer for in situ polymerization in CE capillaries and the coating formed on the inner surface has potential chiral selectivity toward various pharmaceuticals, which is equal or better than traditional chiral CE.
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Li, Yin. "Development of Capillary Electrophoresis for Metabolite Profiling". Thesis, University of Nottingham, 2007. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.519419.
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Hsu, Pei-Chun (Lisa). "Capillary electrophoresis improving clinical measurement of clozapine". Thesis, University of Canterbury. School of Biological Sciences, 2008. http://hdl.handle.net/10092/2260.
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Schizophrenia is a mental disorder affecting approximately one percent of the population worldwide. The introduction of the second generation antipsychotic drug, atypical antipsychotic, clozapine, has demonstrated 80% reduction in suicide incident. This drug showed effectiveness in the treatment of resistant schizophrenia, however, high concentrations of clozapine and N-desmethylclozapine in plasma exhibit the development of agranulocytosis, a possible lethal blood disorder. Therefore, constant therapeutic drug monitoring is important for patients who receive clozapine. High performance liquid chromatography (HPLC) is the current assay for clinical clozapine measurement. A different assay, the capillary electrophoresis (CE) was explored in this study. It was found the use of a background electrolyte (BGE) concentration of 60 mM, pH at 2.5, temperature at 22 ℃, voltage applied at 10 kV and sample injection at 23 kV for 1.5 seconds is the optimal condition for clozapine separation using a fused-silica capillary 75 μm in internal diameter (i.d.). The use of 75 μm (i.d.) fused-silica capillary not only permits a larger sample size, but also provided longer detection pathlength which increased the limit of detection for CE. One hundred and eight patients’ samples were analysed by CE and compared with HPLC results obtained from the Canterbury Health Laboratory. A linear regression line of 1.100 was obtained. Seven External Quality Control (EQC) samples were also analysed and compared to the HPLC results gained from the EQC program world wide. A linear regression line of 1.008 and 1.043 were obtained from clozapine and N-desmethylclozapine separation respectively. The developed CE method has shown to be a valid assay for clozapine and N-desmethylclozapine separation and a more cost effective method compared to HPLC.
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Stocking, Christopher Jon. "Capillary electrophoresis and its application to nephrology". Thesis, Birkbeck (University of London), 2006. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.435912.
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