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Title: A thesis on a comparison of methods for determining the moment of inertia of irregular bodies
Creator: Simmons, George E.
Date: 1894
Collection: Electronic Theses & Dissertations

Title: Comparison of alcoholic and acetic fermentation
Creator: Baker, T. F.
Date: 1913
Collection: Electronic Theses & Dissertations

Title: The feasibility of using a sprinkling system as an auxiliary to a heating system
Creator: Auten, Claude I.
Date: 1913
Collection: Electronic Theses & Dissertations

Title: A study of the original acid reaction of milk to phenolphthalein
Creator: Kurtz, M. B.
Date: 1914
Collection: Electronic Theses & Dissertations

Title: Studies on the pathogenesis of the avian tubercle bacteria
Creator: Mosher, Lawrence A.
Date: 1915
Collection: Electronic Theses & Dissertations

Title: Tests for incipient putrefaction of meat
Creator: Weaver, Ralph H.
Date: 1926
Collection: Electronic Theses & Dissertations

Title: Some biochemical studies on seed viability
Creator: Miller, Erston V. (Erston Vinton), 1898-1985
Date: 1926
Collection: Electronic Theses & Dissertations

Title: Silica gel
Creator: Smits, Benjamin Levi
Date: 1926
Collection: Electronic Theses & Dissertations

Title: Teaching applied electricity
Creator: Willoughby, George Alonzo, 1894-
Date: 1926
Collection: Electronic Theses & Dissertations

Title: Porous titanium dioxide nanomaterials for photocatalytic and photovoltaic applications
Creator: Li, Yan (Graduate of Michigan State University)
Date: 2016
Collection: Electronic Theses & Dissertations
Description: "The huge demand for fossil fuels and the risks of the environmental crisis have spurred an interest in renewable energies. Using the clean and abundant solar energy, semiconductor photocatalysis and photovoltaics have evoked tremendous interest. Titanium dioxide (titania, TiO2) has been the focus of the research trend because of its excellent crystallinity, photoreactivity, chemical and thermal stability, and low cost. The research objective presented in this dissertation is to fabricate...
Show more"The huge demand for fossil fuels and the risks of the environmental crisis have spurred an interest in renewable energies. Using the clean and abundant solar energy, semiconductor photocatalysis and photovoltaics have evoked tremendous interest. Titanium dioxide (titania, TiO2) has been the focus of the research trend because of its excellent crystallinity, photoreactivity, chemical and thermal stability, and low cost. The research objective presented in this dissertation is to fabricate titania nanomaterials with tunable porosities, large surface area, unique morphologies, and enhanced capacities of adsorption, electron transport, diffusion, and then apply them in photocatalysis and photovoltaics. A modified non-hydrolytic sol-gel system with calcium carbonate templating was developed to create macro/micro/nano porous anatase titanium dioxide. The hydrolysis rate was lowered by chelating ligands of valeric acid for slow and sufficient precursor coating. CaCO3 was completely removed via acidification, resulting in titania powders with a surface area ranging from 197 to 239 m2/g. The templated TiO2 with a surface area of 239 m2/g and pore diameters of 6-109 nm showed a promising 27% photocatalytic improvement compared to commercial particles, and a 180% increase compared to template-free TiO2. This increase is attributed to the increased catalyst loading capacity and active photocatalytic sites. From hard templates to soft biodegradable natural templates, the rate-controlled sol-gel method was combined with homogenized micro/nano-fibrillated cellulose (MFCs) with an average diameter below 50 nm. Cellulose was removed completely by thermal treatment, and an in-situ coating technique created thin titania films on substrates with a porous structure. The degradation efficiency of the photocatalytic films was related to film thickness and to the Ti(IV)-to-cellulose ratio. Photocatalyst on film eliminated the post separation treatment related to powder catalyst and simplified the purification process. The sol-gel/MFCs precursor was also coated in-situ as the photoanode for dye-sensitized solar cells. It was found that the thickness of the anode film was a dominant factor to the overall performance and efficiency. The 6-layer cell showed a 400303% increase in solar-to-electricity efficiency (1.75%) compared to commercial paste at the same thickness under a simulated solar light irradiation of 100 mW cm-2 (AM 1.5). From non-hydrolytic to hydrolytic, a modified liquid phase deposition (LPD) approach was combined with MFCs. An optimized solvent composition of isopropanol/water ratio of 4 to 1 was found to yield coatings with uniform spherical TiO2 possessing a chain-like morphology oriented along the axis of the decomposed cellulose fibers. The average rate constant and degradation percentage were 0.72±0.09 min-1, 95% for TiF4-cellulose-4IPA1Water films, which increased by 1.88 times over the film prepared without cellulose templates due to the beneficial surface area, pore size, and the unique morphology. The three-dimensional web structure with pseudo one-dimensional sphere-chain could retard the recombination of photogenerated electron-hole pairs and improve the charge transport."--Pages ii-iii.
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Title: Guilt, empathy, and compliance in a naturalistic moral scenario : predicting prosocial and externalizing behavior in 3-7-year-old children
Creator: Listro, Caitlin J.
Date: 2017
Collection: Electronic Theses & Dissertations
Description: Current science offers only limited answers concerning the development of empathy disturbances. Indeed, few studies have attempted to empirically identify the developmental trajectory of empathy to define either normal or aberrant developmental patterns. The present study aimed to use an observational approach to assess empathy, guilt, and obedience in children, and to validate this approach by exploring how these observational measures of child moral behavior associated with characteristics...
Show moreCurrent science offers only limited answers concerning the development of empathy disturbances. Indeed, few studies have attempted to empirically identify the developmental trajectory of empathy to define either normal or aberrant developmental patterns. The present study aimed to use an observational approach to assess empathy, guilt, and obedience in children, and to validate this approach by exploring how these observational measures of child moral behavior associated with characteristics theoretically linked to moral development and antisocial behavior. We utilized a videotaped Picture Tearing task in which the child is presented with a moral dilemma. Trained coders rated the tasks for several child behaviors (e.g. guilt, gaze avoidance, defiance) using a coding scheme adapted from the Lab-TAB (Goldsmith et al., 1993). Variations in moral behavior were investigated using person-centered (cluster analysis) and variable-centered (factor analysis) methods, then associations between resulting behaviors and other relevant child characteristics (temperament, externalizing behaviors) were examined concurrently and over time. In general, results indicated that empathic verbalizations and defiance were consistently associated with externalizing pathology. This association was observed concurrently; empathy did not predict externalizing over time. Overall, these results suggest that compliant without complaint is the most adaptive response at this age. Furthermore, the Picture Tearing task does provide useful data about empathic behavior and its associations in young children. Recommendations are made for adaptations to the task and coding scheme to improve the measurement of moral behavior in future research.
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Title: Ferroelectret nanogenerator (FENG) for mechanical energy harvesting and self-powered flexible electronics
Creator: Li, Wei (Mechanical engineer)
Date: 2017
Collection: Electronic Theses & Dissertations
Description: Harvesting energy from our natural environment has been the focus of multiple research efforts in the past decades. Progress in this field has far-reaching implications for the growing environment problems resulting from greenhouse gas emission of fossil fuels. Furthermore, advances in portable energy scavenging devices will shed light on the development of self-powered and autonomous electronics; which will impact a broad range of applications in wireless sensors, biomedical implants,...
Show moreHarvesting energy from our natural environment has been the focus of multiple research efforts in the past decades. Progress in this field has far-reaching implications for the growing environment problems resulting from greenhouse gas emission of fossil fuels. Furthermore, advances in portable energy scavenging devices will shed light on the development of self-powered and autonomous electronics; which will impact a broad range of applications in wireless sensors, biomedical implants, infrastructure monitoring, and portable/wearable electronics. This thesis research explores the designs, fabrications, simulations, characterizations and applications of flexible thin film nanogenerator based energy harvesting technologies. Materials and designs for flexible nanogenerator based on nanocrystalline aluminum nitride (AlN) thin film are reported. AlN nanoparticles were grown on aluminum layer by pulsed laser deposition (PLD) at room temperature. Piezoresponse force microscopy (PFM) indicates that their electromechanical energy conversion metrics are as high as highly c-axis oriented AlN or ZnO thin film. Polyimide thin film encapsulated the entire structure of flexible nanogenerator to further improve mechanical robustness, protecting the device from invasive chemicals and enhance its potential biocompaibility. Besides, this thesis research introduces polypropylene ferroelectret (PPFE) as the active material in an efficient, flexible, and biocompatible ferroelectret nanogenerator (FENG) device. PPFE is a type of charged polymers with empty voids and inorganic particles that create giant dipoles across the material's thickness. The mechanical-electrical energy conversion mechanism in PPFE films is verified by finite element method (FEM). Investigation of the developed device shows that the magnitudes of the generated voltage and current signals are doubled each time the device is folded, and an increase with magnitude or frequency of the mechanical input is observed. The developed FENGs is sufficient to light twenty commercial green and blue light-emitting diodes (LEDs), and realize a self-powered liquid-crystal display (LCD) that harvests energy from user's touch. A self-powered flexible/foldable keyboard is also demonstrated. Furthermore, this thesis reports the device's intrinsic properties which allow for the bi-directional conversion of energy between electrical and mechanical domains; thus extending its potential use in wearable electronics beyond the power generation realm. This electromechanical coupling, combined with their flexibility and thin film-like form, bestows dual-functional transducing capabilities to the device that are used in this research to demonstrate its use as a thin, wearable, and self-powered loudspeaker or microphone patch. To determine the device's performance and applicability, sound pressure level is characterized in both space and frequency domains for three different configurations. The confirmed device's high performance is further validated through its integration in three proposed systems: a music-playing flag, a sound recording film, and a flexible microphone for security applications.
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Title: Self-directed learning through the eyes of a Buddhist meditator : (non)control, (non)becoming, and (non)judgement
Creator: Nguyen, Cuong Huy
Date: 2017
Collection: Electronic Theses & Dissertations
Description: By using the meditative inquiry approach, I compare and contrast the philosophy underlying Buddhist meditation, based on my personal experiences with and scholarship on the mindfulness practice, with self-directed learning within the tradition of progressive education, based on the case study of the Jefferson County Open School, a progressive school in Lakewood (CO, USA). I found that Buddhist meditation and self-directed learning have a lot in common. In particular, they both emphasize (non...
Show moreBy using the meditative inquiry approach, I compare and contrast the philosophy underlying Buddhist meditation, based on my personal experiences with and scholarship on the mindfulness practice, with self-directed learning within the tradition of progressive education, based on the case study of the Jefferson County Open School, a progressive school in Lakewood (CO, USA). I found that Buddhist meditation and self-directed learning have a lot in common. In particular, they both emphasize (non)control, (non)becoming, and (non)judgement. As a meditative inquiry, this study has brought about a lot of profound inner transformation in terms of both spirituality and philosophy of education, which are explicitly presented in this dissertation.
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Title: Phase sensitive measurements of ferromagnetic Josephson junctions for cryogenic memory applications
Creator: Niedzielski, Bethany Maria
Date: 2017
Collection: Electronic Theses & Dissertations
Description: A Josephson junction is made up of two superconducting layers separated by a barrier. The original Josephson junctions, studied in the early 1960's, contained an insulating barrier. Soon thereafter, junctions with normal-metal barriers were also studied. Ferromagnetic materials were not even theoretically considered as a barrier layer until around 1980, due to the competing order between ferromagnetic and superconducting systems. However, many exciting physical phenomena arise in hybrid...
Show moreA Josephson junction is made up of two superconducting layers separated by a barrier. The original Josephson junctions, studied in the early 1960's, contained an insulating barrier. Soon thereafter, junctions with normal-metal barriers were also studied. Ferromagnetic materials were not even theoretically considered as a barrier layer until around 1980, due to the competing order between ferromagnetic and superconducting systems. However, many exciting physical phenomena arise in hybrid superconductor/ferromagnetic devices, including devices where the ground state phase difference between the two superconductors is shifted by π. Since their experimental debut in 2001, so-called π junctions have been demonstrated by many groups, including my own, in systems with a single ferromagnetic layer. In this type of system, the phase of the junction can be set to either 0 or π depending on the thickness of the ferromagnetic layer. Of interest, however, is the ability to control the phase of a single junction between the 0 and π states. This was theoretically shown to be possible in a system containing two ferromagnetic layers (spin-valve junctions). If the materials and their thicknesses are properly chosen to manipulate the electron pair correlation function, then the phase state of a spin-valve Josephson junction should be capable of switching between the 0 and π phase states when the magnetization directions of the two ferromagnetic layers are oriented in the antiparallel and parallel configurations, respectively. Such a phase-controllable junction would have immediate applications in cryogenic memory, which is a necessary component to an ultra-low power superconducting computer. A fully superconducting computer is estimated to be orders of magnitude more energy-efficient than current semiconductor-based supercomputers.The goal of this work was to experimentally verify this prediction for a phase-controllable ferromagnetic Josephson junction. To address this complicated system, first, studies of junctions with only a single ferromagnetic junction were required to determine the 0-π transition thickness of that material, the decay of the critical current through the junction with thickness, and the switching field of the material. The materials studied included NiFeMo, NiFe, Ni, and NiFeCo. Additionally, roughness studies of several different superconducting base electrodes and normal metal buffer and spacer layers were performed to determine the optimum junction layers. The ferromagnetic layers used were on the order of 1-2 nm thick, so a smooth growth template is imperative to maintain continuous films with in-plane magnetizations. Lastly, single junction spin-valve samples were studied. We are not equipped to measure the phase of a single junction, but series of samples where one ferromagnetic layer is systematically varied in thickness can inform the proper thicknesses needed for 0-π switching based on relative critical current values between the parallel and antiparallel magnetic configurations. Utilizing this background information, two spin-valve samples were incorporated in a superconducting loop so that the relative phase of the two junctions could be investigated. Through this process, the first phase-controllable ferromagnetic Josephson junctions were experimentally demonstrated using phase-sensitive measurement techniques. This provided the proof of concept for the Josephson Magnetic Random Access Memory (JMRAM), a superconducting memory system in development at Northrop Grumman, with whom we collaborate on this work. Phase-controllable systems were successfully demonstrated using two different magnetic material stacks and verified with several analysis techniques.
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Title: Negotiating fairness : a feminist political ecology of fair trade and organic coffee production in Minas Gerais, Brazil
Creator: Meuninck, Rebecca (Rebecca Mari)
Date: 2017
Collection: Electronic Theses & Dissertations
Description: This dissertation uses a feminist political ecology approach to explore the "fairness" of Fair Trade certification. I do this by examining the gendered social, economic, and environmental impacts of Fair Trade at COOPFAM, a Fair Trade and organic certified coffee cooperative in Minas Gerais, Brazil. Fair Trade is a third-party certification system that attempts to address social and economic inequalities facing small-scale coffee farmers, through floor prices and social development premiums....
Show moreThis dissertation uses a feminist political ecology approach to explore the "fairness" of Fair Trade certification. I do this by examining the gendered social, economic, and environmental impacts of Fair Trade at COOPFAM, a Fair Trade and organic certified coffee cooperative in Minas Gerais, Brazil. Fair Trade is a third-party certification system that attempts to address social and economic inequalities facing small-scale coffee farmers, through floor prices and social development premiums. In return, it requires equitable labor practices, adherence to environmental standards, and freedom of association and democratic decision-making within cooperatives. Fair Trade has grown over the past two decades, due to the liberalization of global coffee markets, and an evolution of consumers' desires which favor more socially and environmentally just coffees. In light of this growth, it is critical to ask, how "fair" is Fair Trade, is it equally fair for all farmers, and how do farmers perceive "fairness"? I use multi-sited ethnographic techniques to explore the "fairness" of Fair Trade; I followed the "thing" (coffee), and "the discourse" (the negotiation of fairness) along COOPFAM's international supply chain (Marcus 1995). I conducted interviews with and participant observation among Brazilian Fair Trade farmers and cooperative administrators, as well as foreign coffee buyers, Fair Trade activists, and certifiers. I then analyze the power dynamics in the Fair Trade system at the local level in homes, at the meso level at the cooperative, and at the macro level with their international partners. I argue that Fair Trade is advantageous for COOPFAM and her farmers, because of the assets they leveraged to overcome the common barriers that have stymied other cooperatives and farmers from obtaining Fair Trade and organic certification. Through Fair Trade, COOPFAM farmers enjoy access to international markets and networks of actors in the supply chain that connect the cooperative with social and economic programs. Moreover, Fair Trade provides an economic safety net for the farmers and the cooperative to experiment with novel production practices, technologies, and emerging certification systems. However, through an examination of COOPFAM's experimentation with new certification systems, the challenges of applying global standards to coffee production surface. Standards, first created to meet the needs of farmers in one locale and the desires of consumers in foreign lands, do not always translate well to other cultures and modes of production. Farmers and cooperatives negotiate these standards with buyers and certifiers, but they are on unequal footing. By examining Fair Trade through a gendered lens, we can see that the system is fairer for some farmers than it is for others. Poorer farmers, those who live far away from the cooperative, unmarried women, and widows are not as well served by the cooperative and may struggle to produce enough high-quality coffee to support their families. My exploration of farmers' livelihood strategies shows that Fair Trade coffee production alone is not sufficient to sustain farming families. Rather, COOPFAM's success and the sustainability of Fair Trade as a production system are reliant on farming families' diversity of livelihood strategies and continual innovation to improve coffee quality.
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Title: Holding hands with death : ethical promises and political failures of our humanitarian present
Creator: Ivanovic, Mlado
Date: 2017
Collection: Electronic Theses & Dissertations
Description: "Taking the point of departure in the urgent social challenges tied with current failures of humanitarian management and inclusion of Non-Western others in "developed" Western societies, this dissertation explores the ways in which representation and knowledge about human suffering guides our (un)willingness to act ethically with respect to vulnerable strangers and the difficult conditions they endure." -- Abstract.

Title: Agency in context : a phenomenological study of Chinese college lLearners' intercultural engagement with expatriate instructors
Creator: Larson, Jay B.
Date: 2017
Collection: Electronic Theses & Dissertations
Description: "As China increasingly internationalizes its higher education system, growing numbers of Chinese learners and expatriate instructors meet in the classroom, engaging one another from their disparate cultural and pedagogical standpoints. Despite its widespread occurrence, the phenomenon of Chinese learners and Western instructors engaging one another in pedagogically and culturally Chinese institutions is largely neglected in research. Scholarly literature on Chinese learners is dominated by...
Show more"As China increasingly internationalizes its higher education system, growing numbers of Chinese learners and expatriate instructors meet in the classroom, engaging one another from their disparate cultural and pedagogical standpoints. Despite its widespread occurrence, the phenomenon of Chinese learners and Western instructors engaging one another in pedagogically and culturally Chinese institutions is largely neglected in research. Scholarly literature on Chinese learners is dominated by studies of their experiences as international students on Western campuses. In Chinese environments, authors most often examine perspectives of expatriate instructors on their cultural adjustments. The voice of the Chinese learner in China is rarely heard in research. In this study, I turned a phenomenological lens toward 17 Chinese learners' lived experiences of intercultural classroom engagement at China's Southwest University. Drawing on works of Hall (1997a, 1997b), Said (1978), and Simmel (1971), I devised a Progression Model of Intercultural Engagement to frame participants' evolving perceptions of expatriate instructors as culturally foreign Others. I focused my analysis of learners' perceptions through three overlapping areas of inquiry: (a) learners' sense-making processes and management of intercultural teaching and learning; (b) their perceptions of the expatriate instructor as a physical, social, and cultural presence; and, (c) perceptions of expatriate instructor's course design and teaching. I collected data over a two-month period at Southwest University through methods including classroom observations, Chinese language participant essays, and English language interviews. Findings revealed learners exercised ownership and agency in interpreting and managing intercultural engagement with their instructors. Participants expressed ownership of a perceived physically, linguistically, and academically Chinese environment. Within that environment, they initially characterized expatriate instructor by foreign-ness. Learners made further sense of intercultural experience by situating their instructors' foreign-ness in constructed social, pedagogical, linguistic, and cultural roles. These roles assigned purpose to the expatriate instructor's foreign presence, and framed the meanings learners constructed from intercultural engagement. Finally, participants interpreted their relationships with written and spoken English through engagement with expatriate instructors perceived as social, pedagogical, and cultural embodiments of language. In sum, findings indicate perceived cultural context of the environment, and senses of ownership and agency learners exercised in this environment. These findings contribute to the academic dialogue on intercultural teaching and learning, not only in Chinese institutional contexts, but anywhere knowledge construction must bridge cultural assumptions, epistemologies, and pedagogies. This study can inform further inquiry into international learners on Western campuses, multinational classes in education hubs, and Western learners studying abroad. I address these applications, and others, as I conclude this dissertation with recommendations for practice, and implications for research and theory."--Pages ii-iii.
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Title: Dynamic network analysis with applications to functional neural connectivity
Creator: Golibagh Mahyari, Arash
Date: 2017
Collection: Electronic Theses & Dissertations
Description: "Contemporary neuroimaging techniques provide neural activity recordings with increasing spatial and temporal resolution yielding rich multichannel datasets that can be exploited for detailed description of anatomical and functional connectivity patterns in the brain. Studies indicate that the changes in functional connectivity patterns across spatial and temporal scales play an important role in a wide range of cognitive and executive processes such as memory and attention as well as in the...
Show more"Contemporary neuroimaging techniques provide neural activity recordings with increasing spatial and temporal resolution yielding rich multichannel datasets that can be exploited for detailed description of anatomical and functional connectivity patterns in the brain. Studies indicate that the changes in functional connectivity patterns across spatial and temporal scales play an important role in a wide range of cognitive and executive processes such as memory and attention as well as in the understanding the causes of many neural diseases and psychopathologies such as epilepsy, Alzheimers, Parkinsons and schizophrenia. Early work in the area was limited to the analysis of static brain networks obtained through averaging long-term functional connectivity, thus neglecting possible time-varying connections. There is growing evidence that functional networks dynamically reorganize and coordinate on millisecond scale for the execution of mental processes. Functional networks consist of distinct network states, where each state is defined as a period of time during which the network topology is quasi-stationary. For this reason, there has been an interest in characterizing the dynamics of functional networks using high temporal resolution electroencephalogram recordings. In this thesis, dynamic functional connectivity networks are represented by multiway arrays, tensors, which are able to capture the complete topological structure of the networks. This thesis proposes new methods for both tracking the changes in these dynamic networks and characterizing or summarizing the network states. In order to achieve this goal, a Tucker decomposition based approach is introduced for detecting the change points for task-based electroencephalogram (EEG) functional connectivity networks through calculating the subspace distance between consecutive time steps. This is followed by a tensor-matrix projection based approach for summarizing multiple networks within a time interval. Tensor based summarization approaches do not necessarily result in sparse network and succinct states. Moreover, subspace based summarizations tend to capture the background brain activity more than the low energy sparse activations. For this reason, we propose utilizing the sparse common component and innovations (SCCI) model which simultaneously finds the sparse common component of multiple signals. However, as the number of signals in the model increases, this becomes computationally prohibitive. In this thesis, a hierarchical algorithm to recover the common component in the SCCI model is proposed for large number of signals. The hierarchical recovery of SCCI model solves the time and memory limitations at the expense of a slight decrease in the accuracy. This hierarchical model is used to separate the common and innovation components of functional connectivity networks across time. The innovation components are tracked over time to detect the change points, and the common component of the detected network states are used to obtain the network summarization. SCCI recovery algorithm finds the sparse representation of the common and innovation components of signals with respect to pre-determined dictionaries. However, input signals are not always well-represented by pre-determined dictionaries. In this thesis, a structured dictionary learning algorithm for SCCI model is developed. The proposed method is applied to EEG data collected during a study of error monitoring where two different types of brain responses are elicited in response to the stimulus. The learned dictionaries can discriminate between the response types and extract the error-related potentials (ERP) corresponding to the two responses."--Pages ii-iii.
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Title: Protease-containing membranes for rapid, controlled antibody digestion prior to mass spectrometry analysis
Creator: Pang, Yongle
Date: 2017
Collection: Electronic Theses & Dissertations
Description: "Monoclonal antibodies are the fastest growing class of therapeutic drugs because of their high specificities to targeQt cells. Facile analysis of therapeutic mAbs and their post-translational modifications (PTMs) is essential for quality control, and mass spectrometry (MS) is the most powerful tool for antibody characterization. Conventional antibody characterization workflows contain an in-solution digestion step, which is labor-intensive and time-consuming. Protease-containing membranes...
Show more"Monoclonal antibodies are the fastest growing class of therapeutic drugs because of their high specificities to targeQt cells. Facile analysis of therapeutic mAbs and their post-translational modifications (PTMs) is essential for quality control, and mass spectrometry (MS) is the most powerful tool for antibody characterization. Conventional antibody characterization workflows contain an in-solution digestion step, which is labor-intensive and time-consuming. Protease-containing membranes are an attractive alternative platform for protein digestion because of their high local enzyme concentrations, short radial diffusion distances, rapid convection in pores, simple fabrication and low cost. Additionally, variation of protein residence time in the membrane gives control over the size of proteolytic peptides. This research focuses on developing workflows for monoclonal antibody characterization using functionalized porous membranes. Sequential adsorption of poly (styrene sulfonate) and pepsin in a porous nylon membrane forms a pepsin membrane reactor. Pepsin is inexpensive and catalyzes proteolysis in acidic solutions, which avoids the need to alkylate cysteine residues and limits antibody deamidation. Variation of the residence times (3 ms to 3 s) of antibody solutions in pepsin-containing membranes yields "bottom-up" (1-2 kDa) to 'middle-down' (5-15 kDa) peptides in less than 10 min. These peptic peptides cover the entire sequences of Herceptin and a WatersTM antibody. Compared with the performance of bottom-up (in-solution tryptic digestion) and top-down (intact protein fragmentation) analysis of an antibody light chain, middle-down (in-membrane peptic digestion) analysis gives the highest bond cleavage (99%). In-membrane digestion also facilitates detection of PTMs such as oxidation, deamidation, N-terminal pyroglutamic acid formation and glycosylation. Recently developed protease-containing spin membranes provide an excellent platform for rapid, membrane-based protein digestion prior to ultrahigh-resolution Orbitrap MS analysis. Centrifugation of 100-200 æL of pretreated protein solutions through the pepsin- or trypsin-containing membranes takes less than 1 min and gives nearly 100% coverage of the protein sequences in subsequent direct infusion MS analysis of digests of apomyoglobin and four commercial monoclonal antibodies (Herceptin, Avastin, Rituxan and Vectibix). MS analysis of peptic and tryptic peptides also reveals mAb PTMs such as N-terminal pyroglutamate formation, C-terminal Lysine clipping and glycosylation. Liquid chromatography coupled to tandem mass spectrometry analysis of tryptic spin digests and subsequent MaxQuant data searching show 100% sequence coverage of all four antibody light chains, and 75.1%-98.4% coverage of the heavy chains. Compared to in-solution tryptic digestion of mAbs, spin digestion yields higher sequence coverage and a larger number of unique peptides. In-membrane digestion also facilitates protein sequence comparison. Rapid peptic in-membrane digestion of two antibodies with direct infusion MS analysis accurately reveals the antibody modification site in less than 1 h. Overall, membrane-based protein digestion uses minimal sample preparation time and yields high peptide and sequence coverages for identification of protein PTMs."--Page ii-iii.
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Title: Dual role of PU.1 in enhancer priming in macrophages
Creator: Tagore, Mohita Malay
Date: 2017
Collection: Electronic Theses & Dissertations
Description: All multicellular organisms arise from a single-celled zygote by the precise execution of a gene expression program which ensures appropriate cell identity. This process is particularly challenging in eukaryotic cells since eukaryotic DNA is packaged by architectural proteins called histones into chromatin, which might act as a barrier to the transcriptional machinery. Macrophages are cells of the immune system which undergo rapid, large scale changes in gene expression in response to...
Show moreAll multicellular organisms arise from a single-celled zygote by the precise execution of a gene expression program which ensures appropriate cell identity. This process is particularly challenging in eukaryotic cells since eukaryotic DNA is packaged by architectural proteins called histones into chromatin, which might act as a barrier to the transcriptional machinery. Macrophages are cells of the immune system which undergo rapid, large scale changes in gene expression in response to bacterial or viral challenge. This makes macrophages an excellent model for studying cell-type specific as well as inducible gene expression. Studies at the genome-wide level have shown that distal regulatory elements like enhancers play an essential role in determining the macrophage inducible response to microbial challenge. Further, lineage-specific transcription factors like PU.1 and C/EBPβ are known to bind inducible enhancers prior to gene induction in resting macrophages. Earlier studies using genome-wide approaches indicate that PU.1 is able to interact with chromatin, thus functioning as a 'pioneer factor' in macrophages. However, not much is known about the mechanism by which PU.1 keeps enhancers accessible prior to gene induction in resting macrophages. Using bone-marrow derived primary mouse macrophage cells as well as PU.1 deficient cell lines, my work highlights the changes in chromatin associated with PU.1 binding during macrophage differentiation as well as in response to bacterial infection. Using a quantitative nucleosome occupancy assay, we reported that PU.1 binding correlates with low nucleosome occupancy at an inducible enhancer in resting macrophages. Further upon induction with an appropriate stimulus, nucleosomes are stably evicted from the distal enhancer and the corresponding gene can be induced. More importantly, my results suggest that lack of PU.1 binding renders regulatory regions (enhancers and promoters) of inducible genes susceptible to heterochromatin formation and silencing by Polycomb repressive complex 2 (PRC2) in differentiated macrophages. PRC2-mediated silencing is also associated with an increase in nucleosome occupancy at the target regions and the corresponding genes cannot be induced. Results obtained from this research will provide important insights into the role of lineage-specific transcription factors at regulatory elements both during normal development and disease.
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