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Alzheimer’s disease (AD) is a very complex disorder and the most common form of dementia. The two pathological hallmarks of AD are extracellular amyloid-β (Aβ) plaques in cerebral cortex, and intraneuronal neurofibrillary tangles. In the early stages of the disease it can be difficult to accurately diagnose AD, as it is difficult to distinguish from normal signs of aging. There is thus a need for sensitive non-invasive tools, able to detect pathophysiological biomarker changes. One such approach is molecular imaging of Aβ plaque load in brain, using PET (positron emission tomography) ligands. We have developed and characterized two novel Aβ plaque neuroimaging PET ligands, AZD2184 and AZD4694. The 2-pyridylbenzothiazole derivate AZD2184, is a 11C-labeled PET ligand with a higher signal-to-background ratio compared to the widely used PET ligand PIB, a 11C-labeled phenylbenzothiazole based tool. This makes it possible to detect smaller changes in Aβ plaque deposition load, and therefore theoretically, also earlier diagnosis. A drawback with 11C-labeled PET ligands is the relatively short half-life. To meet the need for PET ligands with a longer half-life, we developed the pyridylbenzofuran derivate [18F]AZD4694. Although development of fluorinated radioligands is challenging due to the lipophilic nature of aromatic fluorine, we successfully developed a 18F-labeled PET ligand with a signal-to-background ratio matching PIB, the most widely used 11C-labeled PET ligand in clinical use. 3H-labeled derivates of AZD2184, AZD4694, and PIB, showed lower binding specificity towards Aβ plaques containing ApoE. The ApoE genotype per se did not significantly affect ligand binding, instead, the amount of ApoE incorporated to the Aβ plaques appears to be of importance for the binding characteristics of these amyloid PET ligands. Beta-secretase 1 (BACE1) mediates the first step in the processing of amyloid precursor protein (APP) to Aβ peptides, making BACE1 inhibition an attractive therapeutic target in AD. We developed and characterized three novel BACE1 inhibitors, AZD3839, AZ-4217, and AZD3293. AZD3839 and AZ-4217 contains an amidine group which interacts with the catalytic aspartases Asp-32 and Asp-228 of BACE1, effectively inhibiting the enzyme. All three compounds are potent and selective inhibitors of human BACE1, with in vitro potency demonstrated in several cellular models, including primary cortical neurons. All three compound exhibited dose- and time-dependent lowering of plasma, brain, and cerebrospinal fluid Aβ levels in several species, and two of the compounds (AZD3839 and AZD3293) were progressed into clinical trials. / <p>At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 3: Submitted.</p>

Label-free imaging of viruses and nanoparticles directly in complex solutions is important for virology, vaccine research, and rapid diagnostics. These fields would all benefit from tools that allow for more rapid and sensitive characterization of viruses. Traditionally, light microscopy has been used in laboratories for detection of parasites, fungi, and bacteria for both research and clinical diagnosis because it is portable and simple to use. However, virus particles typically cannot be explored using light microscopy without the use of secondary labels due to their small size and low contrast. Characterization and detection of virus particles therefore rely on more complex approaches such as electron microscopy, ELISA, or plaque assay. These approaches require a significant level of expertise, purification of the virus from its natural environment, and often offer indirect verification of the virus presence. A successful virus visualization technique should be rapid, sensitive, and inexpensive, while needing minimal sample preparation or user expertise. We have developed a disposable cartridge based platform for real-time, sensitive, and label free visualization of viruses and nanoparticles directly in complex solutions such as serum. To create this platform we combined an interference reflectance imaging technique (SP-IRIS) with a sealable microfluidic cartridge. Through empirical testing and numeric modelling, the cartridge parameters were optimized and a flow rate of ~3 µL/min was established as optimal. A complex 2-dimensional paper based capillary pump was incorporated into the polymer cartridge to achieve a constant flow rate. Using this platform we were able to reliably show virus detection in a 20 minute experiment. We demonstrate sensitivity comparable to laboratory-based assays such as ELISA and plaque assay, and equal or better sensitivity compared to paper based rapid diagnostic tests. These results display a platform technology that is capable of rapid multiplexed detection and visualization of viruses and nanoparticles directly in solution. This disposable cartridge based platform represents a new approach for sample-to-answer label-free detection and visualization of viruses and nanoparticles. This technology has the potential to enable rapid and high-throughput investigation of virus particle morphology, as well as be used as a rapid point-of-care diagnostic tool where imaging viruses directly in biological samples would be valuable.

archives@tulane.edu / This dissertation is rooted in clinical pathology research, and the main character is addressing limitations in current pathology evaluation workflows. Diagnostic procedures for cancer are typically conducted via core needle biopsy procedures; however, tissue sampling limitations often result in a low yield of samples containing cancer – there are no reliable intraoperative methods to determine if the “lesion is in the needle”. If biopsy procedures result in a diagnosis of cancer, surgical removal of the tumor is often the frontline curative therapy for many cancers. Importantly, histologic evaluation following the whole resected organ is necessary to determine the presence of residual cancer, yet current methods do not allow efficient determination of tumor removal completeness intraoperatively. To address limitations of current histopathology methods, what is critically needed is a point-of-procedure fresh tissue evaluation system that facilitates 1) rapid on-site imaging and evaluation, 2) less destruction, and 3) more complete assessment of tumor content in fresh specimens. A novel microscopy system using video-rate structured illumination (VR-SIM), has been developed with the intent of rapid, point-of-procedure histological screening of intact biopsy and whole surgical specimens. VR-SIM leverages widefield imaging, rapid acting fluorescent stains, and optical sectioning to provide high contrast digital images of tissue with histological relevance. The method is to replicate the standard approach as closely as possible, but replace the physical section with an optically sectioned digital image. The overall goal of this work is to perform technological and methodological refinements necessary to translate VR-SIM as a clinical tool for histologic evaluation of fresh tissue in diagnostic procedures, biobanking, and tumor margin assessment. This project will lay the groundwork for quantitative evaluation of VR-SIM as a clinical tool – with the goal of leading toward industrial design of a VR-SIM as a medical device for hospital use. Developing a new framework for integration of high throughput microscopy into the clinical and research workflow, as well as developing new methods for quantification and evaluation of clinical effectiveness these tools will be presented and discussed in the context of patient outcomes and economic impact. / 1 / David Tulman

The study of high-speed flows represents a challenging problem in the fluid dynamics field due to the presence of chemical reactions and non-equilibrium effects. Hypersonic flights, where speeds reach Mach 5 and above, are particularly influenced by these effects, resulting in a direct impact on the flow and consequently on the aerodynamic performance of a vehicle traveling at these speeds. The study of hypersonic flow conditions requires the experimental capability of determining local temperatures, pressures and velocities using non-intrusive techniques. Furthermore, the simultaneous measurement of two or more variables in a complex flow boosts the amount of information that is obtained since valuable correlations can be established. This research includes the design, construction and characterization of a hypersonic flow apparatus explicitly intended as a tool for advanced laser diagnostics development. This apparatus is characterized by its pulsed operation mode that translates into a significant reduction in mass flow rates and can be operated for long periods at Mach numbers ranging from 2.8 to 6.2. The flow conditions during the uniform flow time interval of each pulse vary by less than 1%, generating a flow of sufficient quality for quantitative measurements. The development of a laser diagnostic technique, the VENOM technique, which is a non-intrusive method to provide simultaneous 2-D measurements of the mean and instantaneous fluctuations in two-component velocity and temperature is also presented. This technique represents the first single diagnostic capable of instantaneous two-component velocimetry and thermometry in a gaseous flow field by combining two Nitric Oxide Planar Laser Induced Fluorescence methods: two-component Molecular Tagging Velocimetry and two-line thermometry, employing the nascent NO(v"=1) arising from the NO2 photodissociation as a molecular tracer. The VENOM technique is expected to be not only applicable to cold high-speed flows, which is the focus of the present work, but also to combustion and other reactive or high-enthalpy flow fields.

Lygus Hahn (Hemiptera: Miridae) are serious pests of economically important field, fruit, vegetable, and greenhouse crops in Canada. The release of European Peristenus Förster (Hymenoptera: Braconidae) in the USA has resulted in significant suppression of this pest and has renewed interest in the release of European Peristenus spp. in Canada. Prior to the release of exotic Peristenus spp., ecological host range studies need to be conducted to define their habitat and host associations. <p>These associations can be difficult to study using conventional methods. Morphological similarity of related parasitoids prevents species-level identification by dissection. Host rearing is time-consuming and can result in high levels of host and parasitoid mortality. To facilitate identification of immature Peristenus spp. in their hosts, a multiplex PCR assay was developed. This assay provided a specific and sensitive tool to screen individual insects for three parasitoid species simultaneously. <p>To validate the utility of the multiplex PCR assay in ecological host range studies, parasitism and parasitoid species composition obtained using conventional and molecular techniques were compared. Molecular methods compared favorably with conventional methods; however, more complete species composition information was available with the multiplex assay. To improve the quality of risk assessment studies and extract the most accurate ecological host range data, molecular methods were used to evaluate host-parasitoid associations in mirid populations collected in two ecoregions. Several new host-parasitoid associations were recorded for <i>P. digoneutis</i> and <i>P. relictus</i>, but parasitism of non-target mirids was low. <p>Parasitism of the target host collected from different plant species was evaluated to help clarify Peristenus host-plant associations. Despite the investigation of three different host plant species, no difference was observed in the parasitism level or parasitoid species composition in <i>L. rugulipennis</i>. The post-release utility of the multiplex assay was investigated in Canada, where Lygus parasitoids may have dispersed following release in the USA. To confirm establishment, samples were analyzed using the multiplex PCR assay, and P. digoneutis was detected for the first time in southern Ontario.

Lygus Hahn (Hemiptera: Miridae) are serious pests of economically important field, fruit, vegetable, and greenhouse crops in Canada. The release of European Peristenus Förster (Hymenoptera: Braconidae) in the USA has resulted in significant suppression of this pest and has renewed interest in the release of European Peristenus spp. in Canada. Prior to the release of exotic Peristenus spp., ecological host range studies need to be conducted to define their habitat and host associations. <p>These associations can be difficult to study using conventional methods. Morphological similarity of related parasitoids prevents species-level identification by dissection. Host rearing is time-consuming and can result in high levels of host and parasitoid mortality. To facilitate identification of immature Peristenus spp. in their hosts, a multiplex PCR assay was developed. This assay provided a specific and sensitive tool to screen individual insects for three parasitoid species simultaneously. <p>To validate the utility of the multiplex PCR assay in ecological host range studies, parasitism and parasitoid species composition obtained using conventional and molecular techniques were compared. Molecular methods compared favorably with conventional methods; however, more complete species composition information was available with the multiplex assay. To improve the quality of risk assessment studies and extract the most accurate ecological host range data, molecular methods were used to evaluate host-parasitoid associations in mirid populations collected in two ecoregions. Several new host-parasitoid associations were recorded for <i>P. digoneutis</i> and <i>P. relictus</i>, but parasitism of non-target mirids was low. <p>Parasitism of the target host collected from different plant species was evaluated to help clarify Peristenus host-plant associations. Despite the investigation of three different host plant species, no difference was observed in the parasitism level or parasitoid species composition in <i>L. rugulipennis</i>. The post-release utility of the multiplex assay was investigated in Canada, where Lygus parasitoids may have dispersed following release in the USA. To confirm establishment, samples were analyzed using the multiplex PCR assay, and P. digoneutis was detected for the first time in southern Ontario.

A Multiagent Framework for a Diagnostic and Prognostic System Irtaza Barlas 124 Pages Directed By: Dr. George Vactsevanos The shortcomings of the current diagnostic and prognostic systems stem from the limitations of their frameworks. The framework is typically designed on the passive, open loop, static, and isolated notions of diagnostics, in that the framework does not observe its diagnostic results (open-looped), hence can not improve its performance (static). Its passivity is attributed to the fact that an external event triggers the diagnostic or prognostic action. There is also no effort in place to team-up the diagnostic systems for a collective learning, hence the implementation is isolated. In this research we extend the current approaches of the design and implementation of diagnostic and prognostic systems by presenting a framework based upon Multiagent systems. This research created novel architectures by providing such unique features to the framework, as learning, reasoning, and coordination. As the primary focus of the research the concept of Case-Based Reasoning was exploited to reason in the temporal domain to generate better prognosis, and improve the accuracy of detection as well as prediction. It was shown that the dynamic behavior of the intelligent agent helps it to learn over time, resulting in improved performance. An analysis is presented to show that a coordinated effort to diagnose also makes sense in uncertain situations when there are certain number of systems attempting to communicate certain number of failures, since there can be high probability of finding a shareable experience.

Fast mixing remains a major challenge in droplet-based microfluidics. The low Reynolds number operating regime typical of most microfluidic devices signify laminar and orderly flows that are devoid of any inertial characteristics. To increase mixing rates in droplet-based devices, a novel technique is presented that uses a high Reynolds number gaseous phase for droplet generation and transport and promotes mixing through binary droplet collisions at velocities near 1m/s. Control of multiple gas and liquid streams is provided by a newly constructed microfluidic test bed that affords the stringent flow stability required for generating liquid droplets in gaseous flows. The result is droplet production with size dispersion and generation frequencies not previously achievable. Limitations of existing mixing diagnostic methods have led to the development of a new measurement technique for measuring droplet collision mixing in confined microchannels. The technique employs single fluorophore laser-induced fluorescence, custom image processing, and meaningful statistical analysis for monitoring and quantifying mixing in high-speed droplet collisions. Mixing information is revealed through three governing statistics that that separate the roles of convective rearrangement and molecular diffusion during the mixing process. The end result is a viewing window into the rich dynamics of droplet collisions with spatial and temporal resolutions of 1μm and 25μs, respectively. Experimental results obtained across a decade vi of Reynolds and Peclet numbers reveal a direct link between droplet mixing time and the collision convective timescale. Increasing the collision velocity or reducing the collision length scale is the most direct method for increasing droplet mixing rates. These characteristics are complemented by detaching droplets under inertial conditions, where increasing the Reynolds number of the continuous gaseous phase generates and transports smaller droplets at faster rates. This work provides valuable insight into the emerging field of two-phase gas-liquid microfluidics and opens the door to fundamental research possibilities not offered by traditional oil-based architectures. / text

Η εργασία αυτή εκπονήθηκε προκειμένου να μελετηθεί η διάγνωση λανθασμένης λειτουργίας μίας μηχανής αυτοκινήτου με μία τεχνική η οποία, ακολούθως, αναλύεται και περιγράφεται. Η μηχανή ΄΄στέλνει΄΄ ένα ημιτονοειδές σήμα το οποίο λαμβάνεται με τη βοήθεια πιεζοηλεκτρικών υλικών. Αυτό καθίσταται δυνατό χάρη στην ιδιότητα που έχουν να μετατρέπουν τη μηχανική τάση ή πίεση σε ηλεκτρική. Αφότου γίνει αυτή η μετατροπή αυτό που πρέπει να ακολουθήσει είναι η δειγματοληψία του σήματος που λαμβάνεται και η επεξεργασία του. Έτσι μπορεί να γίνει η διάγνωση της κατάστασης στην οποία βρίσκεται η μηχανή. Πιο συγκεκριμένα, κατά τη διάρκεια αυτής της εργασίας πραγματοποιείται εξέταση των συστημάτων διάγνωσης λανθασμένης λειτουργίας μίας μηχανής, των χαρακτηριστικών και της χρήσης των σύγχρονων ενσωματωμένων συστημάτων και των μικροελεγκτών και, ειδικά, του μικροελεγκτή ADuC7026 της Analog Devices, ο οποίος και χρησιμοποιείται για την υλοποίηση και εκτέλεση του κώδικα που απαιτείται. Η υλοποίηση της εργασίας βασίστηκε εξ ολοκλήρου σε προσομοίωση που πραγματοποιήθηκε (δεν υπήρξαν εξωτερικές συσκευές ανάπτυξης) και για αυτή την υλοποίηση χρησιμοποιήθηκε προσεγγιστικό ημιτονοειδές σήμα. Κατά την επεξεργασία του σήματος χρησιμοποιήθηκαν FIR (ψηφιακά φίλτρα πεπερασμένης κρουστικής απόκρισης) και αλγόριθμοι εντοπισμού της συχνότητάς του. Για την ανάπτυξη της εφαρμογής που αποσκοπεί στον προγραμματισμό του μικροελεγκτή χρησιμοποιήθηκε το μvision 4 της Keil, το οποίο είναι ένα περιβάλλον ανάπτυξης εφαρμογών και η γλώσσα που χρησιμοποιήθηκε είναι η C. / This work was conducted in order to study the diagnosis of a car's machine malfunction with a technique which is subsequently analyzed and described. The machine sends a sinusoidal signal which is obtained by means of piezoelectric materials. This is made possible thanks to the capacity they have to convert mechanical pressure or voltage to electricity. After this conversion is done what should follow is to sample the signal received and processed. This can make the diagnosis of the condition in which the engine is. More specifically, during this work an examination of diagnostic malfunction of a machine is done, its features and the use of modern embedded systems and microcontrollers and, especially, the microcontroller ADuC7026 of Analog Devices, which is used for the implementation and execution of code required. The implementation work was based entirely on simulation performed (there were no external devices) and this implementation approximate sinusoidal signal was used. In signal processing FIR (Finite Impulse digital response) and frequency detection algorithms were used. For the application aimed at programming the microcontroller what was used is μvision 4 from Keil, which is an environment for development of applications and the language that was used was C.