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  • About
  • The Global ETD Search service is a free service for researchers to find electronic theses and dissertations. This service is provided by the Networked Digital Library of Theses and Dissertations.
    Our metadata is collected from universities around the world. If you manage a university/consortium/country archive and want to be added, details can be found on the NDLTD website.
51

Quantitation of sperm distribution into the fractions during a temperature controlled differential extraction procedure

Ruigrok, Erin Kasey 09 June 2023 (has links)
The typical differential extraction procedure utilized by the forensic science community to extract male deoxyribonucleic acid (DNA) from the sperm cells of the perpetrator separately from female DNA from the epithelial cells of the victim is both time-consuming and labor-intensive. This has contributed greatly to the backlog of unanalyzed sexual assault evidence collection kits (SAECK) seen in many laboratories today and has encouraged research in new methods that are more efficient and more effective in achieving better sperm DNA recovery. The Cotton Lab has developed a Temperature-Controlled Differential Extraction (TCDE) procedure geared towards attaining better sperm recovery and better distribution of male DNA in the sperm fraction (SF) to generate a single source or distinguishable male profile. The TCDE protocol is a direct-lysis procedure that utilizes highly temperature-controlled enzymes, or enzymes that are active at or near their optimal temperatures. This procedure has been previously shown to decrease extraction time significantly and to extract samples that are suitable for downstream analysis. This research specifically attempted to modify the TCDE procedure in the hopes of obtaining higher sperm DNA recovery and eliminating previous concerns of too much sperm being retained by the cotton swab material. It also compared a slightly modified TCDE procedure where the material fraction (MF) and SF are kept as separate fractions (the Separate Method) and a method that results in a recombined MF and SF (Recombined Method) to see if there was a greater distribution of the total male DNA eluted into the SF. Preliminary experimentation with swabs prepared with semen was performed to help make effective modifications. Then, vaginal swabs from eight different female donors were prepared with semen to mimic forensic casework samples and extracted using the Separate and Recombined Methods for comparison of the two extraction methods. Despite unusual epithelial cell lysis results for some samples, the quantitation of the fractions by quantitative polymerase chain reaction (qPCR) showed that for approximately half of the samples extracted using the Separate Method, a majority of total male DNA was eluted into the SF. For these samples, a single source or distinguishable male profile can be generated. However, it was also demonstrated that even with good separation, a very small proportion of the female DNA in the SF still overwhelms the male DNA that is present in much smaller amounts, particularly for the Recombined Method where there are only two fractions. Though further experimentation is necessary, these modifications proved effective in achieving high sperm recovery in the SF and generating a distinguishable male profile when extracting samples using the Separate Method. This research has confirmed that the TCDE procedure can be faster and less labor intensive while still producing clean DNA profiles in downstream analysis, and thus has the potential to be implemented in forensic laboratories after some of the concerns are addressed.
52

Bridging Inductively Coupled Plasma - Mass Spectrometry (ICP-MS) from Metalloproteomics to the Undergraduate Curriculum

Donnell, Anna M. 30 October 2017 (has links)
No description available.
53

Microfluidics for Cell Manipulation and Analysis

Loufakis, Despina Nelie 21 October 2014 (has links)
Microfluidic devices are ideal for analysis of biological systems. The small dimensions result to controlled handling of the flow profile and the cells in suspension. Implementation of additional forces in the system, such as an electric field, promote further manipulation of the cells. In this dissertation, I show novel, unique microfluidic approaches for manipulation and analysis of mammalian cells by the aid of electrical methods or the architecture of the device. Specifically, for the first time, it is shown, that adoption of electrical methods, using surface electrodes, promotes cell concentration in a microchamber due to isoelectric focusing (IEF). In contrast to conventional IEF techniques for protein separation, a matrix is not required in our system, the presence of which would even block the movement of the bulky cells. Electric field is, also, used to breach the cell membrane and gain access to the cell interior by electroporation (irreversible and reversible). Irreversible electroporation is used in a unique, integrated microfluidic device for cell lysis and reagentless extraction of DNA. The genomic material is subsequently analyzed by on-chip PCR, demonstrating the possible elimination of the purification step. On the other hand, reversible electroporation is used for the delivery of exogenous molecules to cells. For the first time, the effect of shear stress on the electroporation efficiency of both attached and suspended cells is examined. On the second part of my dissertation, I explore the capabilities of the architecture of microfluidic devices for cell analysis. A simple, unique method for compartmentalization of a microchamber in an array of picochambers is presented. The main idea of the device lies on the fabrication of solid supports on the main layer of the device. These features may even hold a dual nature (e.g. for cell trapping, and chamber support), in which case, single cell analysis is possible (such as single cell PCR). On the final chapter of my dissertation, a computational analysis of the flow and concentration profiles of a device with hydrodynamic focusing is conducted. I anticipate, that all these novel techniques will be used on integrated microfluidic systems for cell analysis, towards point-of-care diagnostics. / Ph. D.
54

Funktionelle Charakterisierung potentieller Pathogenitätsfaktoren aus Pseudomonas aeruginosa mittels biochemischer und evolutiver Methoden / functional characterization of potential pathogenicity factors from Pseudomonas aeruginosa by biochemical and evolutionary methods

Adams, Thorsten 27 January 2005 (has links)
No description available.
55

Dissecting molecular elements of mitophagy and the lysis of intravacuolar vesicles

Montino, Marco 30 June 2015 (has links)
No description available.
56

Modelo de regressão log-Weibull modificado e a nova distribuição Weibull modificada generalizada / Log-modified Weibull regression models and a new generalized modified Weibull distribution

Farfán Carrasco, Jalmar Manuel 09 November 2007 (has links)
Neste trabalho propomos um modelo de regress~ao utilizando a distribuição Weibull modificado, esta distribuição pode ser usada para modelar dados de sobrevivência quando a de função de risco tem forma de U ou banheira. Assumindo dados censurados, é considerado os estimadores de máxima verossimilhança e Jackknife para os parâmetros do modelo proposto. Foram derivadas as matrizes apropriadas para avaliar influiência local sobre os parâmetros estimados considerando diferentes peturbações e também é apresen- tada alguma medidas de influência global. Para diferentes parâmetros fixados, tamanhos de amostra e porcentagem de censuras, varia simulações foram feitas para avaliar a distribuição empírica do resíduo deviance modificado e comparado coma distribuição normal padrão. Esses estudos sugerem que a distribuição empírica do resíduo devianve modificado para o modelo de regressão log-Weibull modificado com dados censurados aproxima-se de uma dis- tribuição normal padrão. Finalmente analisamos um conjunto de dados utilizando o modelo de regressão log-Weibull modificado. Uma nova distribuição de quatro parâmetros é definida para modelar dados de tempo de vida. Algumas propriedades da distribuição é discutida, assim como ilustramos com exemplos a aplicação dessa nova distribuição. Palavras-chaves: Modelo de regressão; Distribuição Weibull modificada; Distribuição weibull modificada generalizada; Análise de sensibilidade; Dados censurados; Análise de resíduo / In this paperwork are proposed a regression model considering the modified Weibull distribution. This distribution can be used to model bathtub-shaped failure rate functions. Assuming censored data, we consider a classic and Jackknife estimator for the parameters of the model. We derive the appropriate matrices for assessing local influence on the parameter estimates under diferent perturbation schemes and we also present some ways to perform global influence. Besides, for diferent parameter settings, sample sizes and censoring percentages, various simulations are performed and the empirical distribution of the deviance modified residual is displayed and compared with the standard normal distribution. These studies suggest that the residual analysis usually performed in normal linear regression models can be straightforwardly extend for a martingale-type residual in log-modifiedWeibull regression models with censored data. Finally, we analyze a real data set under log-modified Weibull regression models. A diagnostic analysis and a model checking based on the deviance modified residual are performed to select an appropriate model. A new four-parameter distribution is introduced. Various properties the new distribution are discussed. Illustrative examples based on real data are also given.
57

Antileukemic activity of allogeneic NK Cells / Potentiel anti-leucémique des cellules NK allogéniques

Nanbakhsh, Arash 20 October 2014 (has links)
Les cellules tueuses naturelles (NK pour Natural Killer) sont une population lymphoïde dotées d’une activité cytotoxique contre les cellules infectées ou les cellules cancéreuses. Les cellules NK ont un potentiel thérapeutique considérable en tant que thérapie cellulaire anti-tumorale, particulièrement dans le cadre des leucémies. Ces approches sont basées sur une amélioration de la production de cellules NK à partir de cellules souches hématopoïétiques (quantitative et qualitative en améliorant leur activité lytique), mais aussi sur une manipulation de la sensibilité des cellules leucémiques à la lyse par les cellules NK. L’amélioration de ces approches nécessite une compréhension plus approfondie des différents mécanismes de résistance leucémique et leur relation avec la sensibilité à la lyse. Dans ce contexte, nous avons étudié le rôle de HOXB4 dans la différenciation des cellules NK et leur fonction lytique. Nous avons montré que les cellules CD34+ différenciées en cellules NK en présence de HOXB4 ont un potentiel lytique plus important par rapport aux cellules différenciées en l’absence de HIOXB4. Cette augmentation est associée à une augmentation de la dégranulation des cellules NK en présence de cellules cibles. L’analyse transcriptionnelle globale basé sur un microréseau d'ADN montre une régulation positive de l’expression de granzyme B par HOXB4. Ces résultats démontrent que HOXB4 est un régulateur crucial dans la différenciation et la fonction des cellules NK. Ils soulignent également l’intérêt de son utilisation dans la production de cellules NK fonctionnelles dotées d’un plus grand potentiel lytique pour les stratégies d'immunothérapie anticancéreuse. Nous avons également essayé de comprendre comment l'acquisition de la résistance aux chimiothérapies par les cellules de leucémie aigüe myéloïde (LAM) influence leur reconnaissance et leur sensibilité aux cellules NK. Nous avons montré que l'acquisition de la résistance in vitro des cellules AML à la cytarabine induit une augmentation de leur susceptibilité à la cytotoxicité dépendante des cellules NK. Cette sensibilité accrue est en corrélation avec l’induction d’ULBP (UL-16 binding proteins) 1/2/3, ligands des récepteurs NKG2D, sur les cellules leucémiques résistantes. Cette induction est régulée par un mécanisme impliquant l'induction de c-Myc. Le test d’immunoprécipitation de la chromatine (ChIP) a révélé qu’ULBP1 et ULBP3 sont des cibles directes de c-Myc. L’utilisation de cellules AML primaires résistants à la chimiothérapie comme les cellules cibles, combinée à l'inhibition de c-Myc a entraîné une diminution de l'expression des ligands NKG2D et l'altération de la lyse par les cellules NK. Les propriétés d’alloréactivité des cellules NK pourraient être utilisées pour améliorer les résultats de la transplantation de cellules souches hématopoïétiques allogéniques chez les patients atteints d’AML. Cependant, la résistance croisée (chimiothérapie et NK) des blastes AML reste un problème majeur. Nous avons étudié la relation entre résistance des cellules leucémiques à la daunorubicine, la susceptibilité de ces cellules à la lyse par les cellules NK et l'expression putative des micro-RNAs. Nos résultats indiquent que l'acquisition de la résistance à la daunorubicine par les lignées de cellules parentales induit une résistance croisée à la cytotoxicité naturelle à médiation cellulaire. L'analyse des microréseaux de microRNAs a révélé que cette résistance croisée est associée à une diminution du miR-181a et une augmentation des gènes de la famille tyrosine kinase (MAP3K10 et MAP2K1) et de la famille Bcl-2 (Bcl-2 et Mcl-1). La surexpression de miR-181a dans les blastes AML entraîne l'atténuation de leur résistance à la daunorobucine et à la lyse par les cellules NK. / Natural Killer (NK) cells are a lymphoid population with potent cytotoxic activity against virus-infected or cancer cells, and which hold considerable potential for cell based therapies targeting human malignancies. Potential approaches include not only enhancing the generation of NK cells in number and improving their lytic activity, but also manipulating the susceptibility of blast cells to NK-mediated killing. Pursuing these approaches will require a more thorough understanding of the different mechanisms of resistance and their relationship with susceptibility to NK-mediated killing. In this context, we studied the role of HOXB4 in NK cells differentiation and lytic function. We showed that HOXB4 transduced MS-5 cells as compared with GFP-transduced MS-5 cells induced highly differentiated cytotoxic NK cells. This difference was associated with an increased induction of granzyme B degranulation in response to stimulation with NK cell susceptible targets. DNA microarray-based global transcriptional profiling confirmed the upregulation of granzyme B. These findings provide further evidence that HOXB4 is a crucial regulator of NK function and that its use in generating functional NK cells with increased lytic potential may be significant for cancer immunotherapy. We attempted to elucidate how acquisition of drug resistance in AML cells influences NK cell recognition and the killing of drug-resistant blasts. We showed that the in vitro acquisition of AML cell resistance to cytarabine resulted in an increase in their susceptibility to NK-mediated cell cytotoxicity. The increased susceptibility correlates with the induction of UL-16 binding proteins (ULBP) 1/2/3 and NK group 2, member (NKG2D) ligands on target cells by a mechanism involving c-Myc induction. More importantly, chromatin immunoprecipitation assay revealed that ULBP1/3 are direct targets of c-Myc. Using drug resistant primary AML blasts as target cells, inhibition of c-Myc resulted in decreased expression of NKG2D ligands and the subsequent impairment of NK cell lysis. This study provides for the first time, the c-Myc dependent regulation of NKG2D ligands in AML. Manipulating NK-cell alloreactivity might improve outcomes after hematopoietic stem-cell transplantation; however, cross-resistance among blasts remains a drawback. We attempted to investigate the relationship between AML to daunorubicin, the susceptibility to NK cellmediated cell lysis and the putative expression of miRs. Our results indicate that the acquisition of resistance to daunorubicin by the parental cell lines resulted in the acquisition of a cross-resistance to natural cell-mediated cytotoxicity. miR microarray analysis revealed that this cross-resistance was associated with miR-181a down regulation and the subsequent regulation of the tyrosine kinase (MAP3K10 and MAP2K1) and the BCL-2 (BCL-2 andMCL-1) families. Overexpression of miR-181a in AML blasts resulted in the attenuation of their resistance to daunorobucin and to NK-cell-mediated killing.
58

Modelo de regressão log-Weibull modificado e a nova distribuição Weibull modificada generalizada / Log-modified Weibull regression models and a new generalized modified Weibull distribution

Jalmar Manuel Farfán Carrasco 09 November 2007 (has links)
Neste trabalho propomos um modelo de regress~ao utilizando a distribuição Weibull modificado, esta distribuição pode ser usada para modelar dados de sobrevivência quando a de função de risco tem forma de U ou banheira. Assumindo dados censurados, é considerado os estimadores de máxima verossimilhança e Jackknife para os parâmetros do modelo proposto. Foram derivadas as matrizes apropriadas para avaliar influiência local sobre os parâmetros estimados considerando diferentes peturbações e também é apresen- tada alguma medidas de influência global. Para diferentes parâmetros fixados, tamanhos de amostra e porcentagem de censuras, varia simulações foram feitas para avaliar a distribuição empírica do resíduo deviance modificado e comparado coma distribuição normal padrão. Esses estudos sugerem que a distribuição empírica do resíduo devianve modificado para o modelo de regressão log-Weibull modificado com dados censurados aproxima-se de uma dis- tribuição normal padrão. Finalmente analisamos um conjunto de dados utilizando o modelo de regressão log-Weibull modificado. Uma nova distribuição de quatro parâmetros é definida para modelar dados de tempo de vida. Algumas propriedades da distribuição é discutida, assim como ilustramos com exemplos a aplicação dessa nova distribuição. Palavras-chaves: Modelo de regressão; Distribuição Weibull modificada; Distribuição weibull modificada generalizada; Análise de sensibilidade; Dados censurados; Análise de resíduo / In this paperwork are proposed a regression model considering the modified Weibull distribution. This distribution can be used to model bathtub-shaped failure rate functions. Assuming censored data, we consider a classic and Jackknife estimator for the parameters of the model. We derive the appropriate matrices for assessing local influence on the parameter estimates under diferent perturbation schemes and we also present some ways to perform global influence. Besides, for diferent parameter settings, sample sizes and censoring percentages, various simulations are performed and the empirical distribution of the deviance modified residual is displayed and compared with the standard normal distribution. These studies suggest that the residual analysis usually performed in normal linear regression models can be straightforwardly extend for a martingale-type residual in log-modifiedWeibull regression models with censored data. Finally, we analyze a real data set under log-modified Weibull regression models. A diagnostic analysis and a model checking based on the deviance modified residual are performed to select an appropriate model. A new four-parameter distribution is introduced. Various properties the new distribution are discussed. Illustrative examples based on real data are also given.
59

Integration of Nanoparticle Cell Lysis and Microchip PCR as a Portable Solution for One-Step Rapid Detection of Bacteria

Wan, Weijie January 2011 (has links)
Bacteria are the oldest, structurally simplest, and most abundant forms of life on earth. Its detection has always been a serious question since the emerging of modern science and technology. There has been a phenomenal growth in the field of real-time bacteria detection in recent years with emerging applications in a wide range of disciplines, including medical analysis, food, environment and many more. Two important analytical functions involved in bacteria detection are cell lysis and polymerase chain reaction (PCR). Cell lysis is required to break cells open to release DNA for use in PCR. PCR is required to reproduce millions of copies of the target genes to reach detection limit from a low DNA concentration. Conventionally, cell lysis and PCR are performed separately using specialized equipments. Those bulky machines consume much more than needed chemical reagents and are very time consuming. An efficient, cost-effective and portable solution involving Nanotechnology and Lab-on-a-Chip (LOC) technology was proposed. The idea was to utilize the excellent antibacterial property of surface-functionalized nanoparticles to perform cell lysis and then to perform PCR on the same LOC system without having to remove them from the solution for rapid detection of bacteria. Nanoparticles possess outstanding properties that are not seen in their bulk form due to their extremely small size. They were introduced to provide two novel methods for LOC cell lysis to overcome problems of current LOC cell lysis methods such as low efficiency, high cost and complicated fabrication process. The first method involved using poly(quaternary ammonium) functionalized gold and titanium dioxide nanoparticles which were demonstrated to be able to lyse E. coli completely in 10 minutes. The idea originated from the excellent antibacterial property of quaternary ammonium salts that people have been using for a long time. The second method involved using titanium dioxide nanoparticles and a miniaturized UV LED array. Titanium dioxide bears photocatalytic effect which generates highly reactive radicals to compromise cell membranes upon absorbing UV light in an aqueous environment. A considerable reduction of live E. coli was observed in 60 minutes. The thesis then evaluates the effect of nanoparticles on PCR to understand the roles nanoparticles play in PCR. It was found that gold and titanium dioxide nanoparticles induce PCR inhibition. How size of gold nanoparticles affected PCR was studied as well. Effective methods were discovered to suppress PCR inhibition caused by gold and titanium dioxide nanoparticles. The pioneering work paves a way for the integration of nanoparticle cell lysis and LOC PCR for rapid detection of bacteria. In the end, an integrated system involving nanoparticle cell lysis and microchip PCR was demonstrated. The prototyped system consisted of a physical microchip for both cell lysis and PCR, a temperature control system and necessary interface connections between the physical device and the temperature control system. The research explored solutions to improve PCR specificity in a microchip environment with gold nanoparticles in PCR. The system was capable of providing the same performance while reducing PCR cycling time by up to 50%. It was inexpensive and easy to be constructed without any complicated clean room fabrication processes. It can find enormous applications in water, food, environment and many more.
60

Integration of Nanoparticle Cell Lysis and Microchip PCR as a Portable Solution for One-Step Rapid Detection of Bacteria

Wan, Weijie January 2011 (has links)
Bacteria are the oldest, structurally simplest, and most abundant forms of life on earth. Its detection has always been a serious question since the emerging of modern science and technology. There has been a phenomenal growth in the field of real-time bacteria detection in recent years with emerging applications in a wide range of disciplines, including medical analysis, food, environment and many more. Two important analytical functions involved in bacteria detection are cell lysis and polymerase chain reaction (PCR). Cell lysis is required to break cells open to release DNA for use in PCR. PCR is required to reproduce millions of copies of the target genes to reach detection limit from a low DNA concentration. Conventionally, cell lysis and PCR are performed separately using specialized equipments. Those bulky machines consume much more than needed chemical reagents and are very time consuming. An efficient, cost-effective and portable solution involving Nanotechnology and Lab-on-a-Chip (LOC) technology was proposed. The idea was to utilize the excellent antibacterial property of surface-functionalized nanoparticles to perform cell lysis and then to perform PCR on the same LOC system without having to remove them from the solution for rapid detection of bacteria. Nanoparticles possess outstanding properties that are not seen in their bulk form due to their extremely small size. They were introduced to provide two novel methods for LOC cell lysis to overcome problems of current LOC cell lysis methods such as low efficiency, high cost and complicated fabrication process. The first method involved using poly(quaternary ammonium) functionalized gold and titanium dioxide nanoparticles which were demonstrated to be able to lyse E. coli completely in 10 minutes. The idea originated from the excellent antibacterial property of quaternary ammonium salts that people have been using for a long time. The second method involved using titanium dioxide nanoparticles and a miniaturized UV LED array. Titanium dioxide bears photocatalytic effect which generates highly reactive radicals to compromise cell membranes upon absorbing UV light in an aqueous environment. A considerable reduction of live E. coli was observed in 60 minutes. The thesis then evaluates the effect of nanoparticles on PCR to understand the roles nanoparticles play in PCR. It was found that gold and titanium dioxide nanoparticles induce PCR inhibition. How size of gold nanoparticles affected PCR was studied as well. Effective methods were discovered to suppress PCR inhibition caused by gold and titanium dioxide nanoparticles. The pioneering work paves a way for the integration of nanoparticle cell lysis and LOC PCR for rapid detection of bacteria. In the end, an integrated system involving nanoparticle cell lysis and microchip PCR was demonstrated. The prototyped system consisted of a physical microchip for both cell lysis and PCR, a temperature control system and necessary interface connections between the physical device and the temperature control system. The research explored solutions to improve PCR specificity in a microchip environment with gold nanoparticles in PCR. The system was capable of providing the same performance while reducing PCR cycling time by up to 50%. It was inexpensive and easy to be constructed without any complicated clean room fabrication processes. It can find enormous applications in water, food, environment and many more.

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