Matching Items (28)
Description
Laboratory automation systems have seen a lot of technological advances in recent times. As a result, the software that is written for them are becoming increasingly sophisticated. Existing software architectures and standards are targeted to a wider domain of software development and need to be customized in order to use them for developing software for laboratory automation systems. This thesis proposes an architecture that is based on existing software architectural paradigms and is specifically tailored to developing software for a laboratory automation system. The architecture is based on fairly autonomous software components that can be distributed across multiple computers. The components in the architecture make use of asynchronous communication methodologies that are facilitated by passing messages between one another. The architecture can be used to develop software that is distributed, responsive and thread-safe. The thesis also proposes a framework that has been developed to implement the ideas proposed by the architecture. The framework is used to develop software that is scalable, distributed, responsive and thread-safe. The framework currently has components to control very commonly used laboratory automation devices such as mechanical stages, cameras, and also to do common laboratory automation functionalities such as imaging.
ContributorsKuppuswamy, Venkataramanan (Author) / Meldrum, Deirdre (Thesis advisor) / Collofello, James (Thesis advisor) / Sarjoughian, Hessam S. (Committee member) / Johnson, Roger (Committee member) / Arizona State University (Publisher)
Created2012
Description
Within the last decade there has been remarkable interest in single-cell metabolic analysis as a key technology for understanding cellular heterogeneity, disease initiation, progression, and drug resistance. Technologies have been developed for oxygen consumption rate (OCR) measurements using various configurations of microfluidic devices. The technical challenges of current approaches include: (1) deposition of multiple sensors for multi-parameter metabolic measurements, e.g. oxygen, pH, etc.; (2) tedious and labor-intensive microwell array fabrication processes; (3) low yield of hermetic sealing between two rigid fused silica parts, even with a compliance layer of PDMS or Parylene-C. In this thesis, several improved microfabrication technologies are developed and demonstrated for analyzing multiple metabolic parameters from single cells, including (1) a modified "lid-on-top" configuration with a multiple sensor trapping (MST) lid which spatially confines multiple sensors to micro-pockets enclosed by lips for hermetic sealing of wells; (2) a multiple step photo-polymerization method for patterning three optical sensors (oxygen, pH and reference) on fused silica and on a polyethylene terephthalate (PET) surface; (3) a photo-polymerization method for patterning tri-color (oxygen, pH and reference) optical sensors on both fused silica and on the PET surface; (4) improved KMPR/SU-8 microfabrication protocols for fabricating microwell arrays that can withstand cell culture conditions. Implementation of these improved microfabrication methods should address the aforementioned challenges and provide a high throughput and multi-parameter single cell metabolic analysis platform.
ContributorsSong, Ganquan (Author) / Meldrum, Deirdre R (Thesis advisor) / Goryll, Michael (Committee member) / Wang, Hong (Committee member) / Tian, Yanqing (Committee member) / Arizona State University (Publisher)
Created2014
Description
Cellular heterogeneity is a key factor in various cellular processes as well as in disease development, especially associated with immune response and cancer progression. Cell-to-cell variability is considered to be one of the major obstacles in early detection and successful treatment of cancer. Most present technologies are based on bulk cell analysis, which results in averaging out the results acquired from a group of cells and hence missing important information about individual cells and their behavior. Understanding the cellular behavior at the single-cell level can help in obtaining a complete profile of the cell and to get a more in-depth knowledge of cellular processes. For example, measuring transmembrane fluxes oxygen can provide a direct readout of the cell metabolism.
The goal of this thesis is to design, optimize and implement a device that can measure the oxygen consumption rate (OCR) of live single cells. A microfluidic device has been designed with the ability to rapidly seal and unseal microchambers containing individual cells and an extracellular optical oxygen sensor for measuring the OCR of live single cells. The device consists of two parts, one with the sensor in microwells (top half) and the other with channels and cells trapped in Pachinko-type micro-traps (bottom half). When the two parts of the device are placed together the wells enclose each cell. Oil is flown in through the channels of the device to produce isolated and sealed microchamber around each cell. Different fluids can be flowed in and out of the device, alternating with oil, to rapidly switch between sealed and unsealed microenvironment around each cell. A fluorescent ratiometric dual pH and oxygen sensor is placed in each well. The thesis focuses on measuring changes in the oxygen consumption rate of each cell within a well. Live and dead cells are identified using a fluorescent live/dead cell assay. Finally, the technology is designed to be scalable for high-throughput applications by controlling the flow rate of the system and increasing the cell array density.
The goal of this thesis is to design, optimize and implement a device that can measure the oxygen consumption rate (OCR) of live single cells. A microfluidic device has been designed with the ability to rapidly seal and unseal microchambers containing individual cells and an extracellular optical oxygen sensor for measuring the OCR of live single cells. The device consists of two parts, one with the sensor in microwells (top half) and the other with channels and cells trapped in Pachinko-type micro-traps (bottom half). When the two parts of the device are placed together the wells enclose each cell. Oil is flown in through the channels of the device to produce isolated and sealed microchamber around each cell. Different fluids can be flowed in and out of the device, alternating with oil, to rapidly switch between sealed and unsealed microenvironment around each cell. A fluorescent ratiometric dual pH and oxygen sensor is placed in each well. The thesis focuses on measuring changes in the oxygen consumption rate of each cell within a well. Live and dead cells are identified using a fluorescent live/dead cell assay. Finally, the technology is designed to be scalable for high-throughput applications by controlling the flow rate of the system and increasing the cell array density.
ContributorsRodrigues, Meryl (Author) / Meldrum, Deirdre (Thesis advisor) / Kelbauskas, Laimonas (Committee member) / LaBelle, Jeffrey (Committee member) / Arizona State University (Publisher)
Created2014
Description
Filtration for microfluidic sample-collection devices is desirable for sample selection, concentration, preprocessing, and downstream manipulation, but microfabricating the required sub-micrometer filtration structure is an elaborate process. This thesis presents a simple method to fabricate polydimethylsiloxane (PDMS) devices with an integrated membrane filter that will sample, lyse, and extract the DNA from microorganisms in aqueous environments. An off-the-shelf membrane filter disc was embedded in a PDMS layer and sequentially bound with other PDMS channel layers. No leakage was observed during filtration. This device was validated by concentrating a large amount of cyanobacterium Synechocystis in simulated sample water with consistent performance across devices. After accumulating sufficient biomass on the filter, a sequential electrochemical lysing process was performed by applying 5VDC across the filter. This device was further evaluated by delivering several samples of differing concentrations of cyanobacterium Synechocystis then quantifying the DNA using real-time PCR. Lastly, an environmental sample was run through the device and the amount of photosynthetic microorganisms present in the water was determined. The major breakthroughs in this design are low energy demand, cheap materials, simple design, straightforward fabrication, and robust performance, together enabling wide-utility of similar chip-based devices for field-deployable operations in environmental micro-biotechnology.
ContributorsLecluse, Aurelie (Author) / Meldrum, Deirdre (Thesis advisor) / Chao, Joseph (Thesis advisor) / Westerhoff, Paul (Committee member) / Arizona State University (Publisher)
Created2011
Description
Single cell phenotypic heterogeneity studies reveal more information about the pathogenesis process than conventional bulk methods. Furthermore, investigation of the individual cellular response mechanism during rapid environmental changes can only be achieved at single cell level. By enabling the study of cellular morphology, a single cell three-dimensional (3D) imaging system can be used to diagnose fatal diseases, such as cancer, at an early stage. One proven method, CellCT, accomplishes 3D imaging by rotating a single cell around a fixed axis. However, some existing cell rotating mechanisms require either intricate microfabrication, and some fail to provide a suitable environment for living cells. This thesis develops a microvorterx chamber that allows living cells to be rotated by hydrodynamic alone while facilitating imaging access. In this thesis work, 1) the new chamber design was developed through numerical simulation. Simulations revealed that in order to form a microvortex in the side chamber, the ratio of the chamber opening to the channel width must be smaller than one. After comparing different chamber designs, the trapezoidal side chamber was selected because it demonstrated controllable circulation and met the imaging requirements. Microvortex properties were not sensitive to the chambers with interface angles ranging from 0.32 to 0.64. A similar trend was observed when chamber heights were larger than chamber opening. 2) Micro-particle image velocimetry was used to characterize microvortices and validate simulation results. Agreement between experimentation and simulation confirmed that numerical simulation was an effective method for chamber design. 3) Finally, cell rotation experiments were performed in the trapezoidal side chamber. The experimental results demonstrated cell rotational rates ranging from 12 to 29 rpm for regular cells. With a volumetric flow rate of 0.5 µL/s, an irregular cell rotated at a mean rate of 97 ± 3 rpm. Rotational rates can be changed by altering inlet flow rates.
ContributorsZhang, Wenjie (Author) / Frakes, David (Thesis advisor) / Meldrum, Deirdre (Thesis advisor) / Chao, Shih-hui (Committee member) / Wang, Xiao (Committee member) / Arizona State University (Publisher)
Created2011
Description
Single cell analysis has become increasingly important in understanding disease onset, progression, treatment and prognosis, especially when applied to cancer where cellular responses are highly heterogeneous. Through the advent of single cell computerized tomography (Cell-CT), researchers and clinicians now have the ability to obtain high resolution three-dimensional (3D) reconstructions of single cells. Yet to date, no live-cell compatible version of the technology exists. In this thesis, a microfluidic chip with the ability to rotate live single cells in hydrodynamic microvortices about an axis parallel to the optical focal plane has been demonstrated. The chip utilizes a novel 3D microchamber design arranged beneath a main channel creating flow detachment into the chamber, producing recirculating flow conditions. Single cells are flowed through the main channel, held in the center of the microvortex by an optical trap, and rotated by the forces induced by the recirculating fluid flow. Computational fluid dynamics (CFD) was employed to optimize the geometry of the microchamber. Two methods for the fabrication of the 3D microchamber were devised: anisotropic etching of silicon and backside diffuser photolithography (BDPL). First, the optimization of the silicon etching conditions was demonstrated through design of experiment (DOE). In addition, a non-conventional method of soft-lithography was demonstrated which incorporates the use of two positive molds, one of the main channel and the other of the microchambers, compressed together during replication to produce a single ultra-thin (<200 µm) negative used for device assembly. Second, methods for using thick negative photoresists such as SU-8 with BDPL have been developed which include a new simple and effective method for promoting the adhesion of SU-8 to glass. An assembly method that bonds two individual ultra-thin (<100 µm) replications of the channel and the microfeatures has also been demonstrated. Finally, a pressure driven pumping system with nanoliter per minute flow rate regulation, sub-second response times, and < 3% flow variability has been designed and characterized. The fabrication and assembly of this device is inexpensive and utilizes simple variants of conventional microfluidic fabrication techniques, making it easily accessible to the single cell analysis community.
ContributorsMyers, Jakrey R (Author) / Meldrum, Deirdre (Thesis advisor) / Johnson, Roger (Committee member) / Frakes, David (Committee member) / Arizona State University (Publisher)
Created2012
Description
Traumatic brain injury (TBI) is a leading cause of disability worldwide with 1.7 million TBIs reported annually in the United States. Broadly, TBI can be classified into focal injury, associated with cerebral contusion, and diffuse injury, a widespread injury pathology. TBI results in a host of pathological alterations and may lead to a transient blood-brain-barrier (BBB) breakdown. Although the BBB dysfunction after TBI may provide a window for therapeutic delivery, the current drug delivery approaches remains largely inefficient due to rapid clearance, inactivation and degradation. One potential strategy to address the current therapeutic limitations is to employ nanoparticle (NP)-based technology to archive greater efficacy and reduced clearance compared to standard drug administration. However, NP application for TBI is challenging not only due to the transient temporal resolution of the BBB breakdown, but also due to the heterogeneous (focal/diffuse) aspect of the disease itself. Furthermore, recent literature suggests sex of the animal influences neuroinflammation/outcome after TBI; yet, the influence of sex on BBB integrity following TBI and subsequent NP delivery has not been previously investigated. The overarching hypothesis for this thesis is that TBI-induced compromised BBB and leaky vasculature will enable delivery of systemically injected NPs to the injury penumbra. This study specifically explored the feasibility and the temporal accumulation of NPs in preclinical mouse models of focal and diffuse TBI. Key findings from these studies include the following. (1) After focal TBI, NPs ranging from 20-500nm exhibited peak accumulation within the injury penumbra acutely (1h) post-injury. (2) A smaller delayed peak of NP accumulation (40nm) was observed sub-acutely (3d) after focal brain injury. (3) Mild diffuse TBI simulated with a mild closed head injury model did not display any measurable NP accumulation after 1h post-injury. (4) In contrast, a moderate diffuse model (fluid percussion injury) demonstrated peak accumulation at 3h post-injury with up to 500 nm size NPs accumulating in cortical tissue. (5) Robust NP accumulation (40nm) was found in female mice compared to the males at 24h and 3d following focal brain injury. Taken together, these results demonstrate the potential for NP delivery at acute and sub-acute time points after TBI by exploiting the compromised BBB. Results also reveal a potential sex dependent component of BBB disruption leading to altered NP accumulation. The applications of this research are far-reaching ranging from theranostic delivery to personalized NP delivery for effective therapeutic outcome.
ContributorsBharadwaj, Vimala Nagabhushana (Author) / Stabenfeldt, Sarah E (Thesis advisor) / Kodibagkar, Vikram D (Thesis advisor) / Kleim, Jeffrey (Committee member) / Tian, Yanqing (Committee member) / Lifshitz, Jonathan (Committee member) / Anderson, Trent R (Committee member) / Arizona State University (Publisher)
Created2018
Description
A series of mitochondria targeting probes was synthesized for the purpose of exploring the feasibility of a mitochondria targeting fluorescent sensor. Of the probes, the probe with a two carbon spacer showed the best co-localization from staining with the established MitoTracker Red® FM, indicating a potential development of the probe into mitochondria targeting sensor. However, cytotoxicity was observed for the probe with a six carbon spacer. Three additional mitochondria targeting fluorescent probes of longer spacer groups were synthesized, but the cytotoxicity was not observed to be as high as that of the probe with a two carbon spacer. The cytotoxicity was characterized to be that of caspase dependent cell death. To screen for a possible effect on apoptosis due to the mitochondrial probe, three fluorescent fusion proteins binding the anti-apoptotic proteins were designed and expressed. Each purified fusion protein was then incubated with the cytotoxic mitochondrial probe, and the mixture was isolated by running an affinity column. The fluorescence analysis of eluted fractions showed preliminary data of possible interaction between the protein and the mitochondrial probe.
ContributorsLee, Fred (Author) / Meldrum, Deirdre R. (Thesis director) / Tian, Yanqing (Committee member) / Zhang, Liqiang (Committee member) / Barrett, The Honors College (Contributor) / Chemical Engineering Program (Contributor) / Department of Chemistry and Biochemistry (Contributor)
Created2014-12
Description
Relapse after tumor dormancy is one of the leading causes of cancer recurrence that ultimately leads to patient mortality. Upon relapse, cancer manifests as metastases that are linked to almost 90% cancer related deaths. Capture of the dormant and relapsed tumor phenotypes in high-throughput will allow for rapid targeted drug discovery, development and validation. Ablation of dormant cancer will not only completely remove the cancer disease, but also will prevent any future recurrence. A novel hydrogel, Amikagel, was developed by crosslinking of aminoglycoside amikacin with a polyethylene glycol crosslinker. Aminoglycosides contain abundant amount of easily conjugable groups such as amino and hydroxyl moieties that were crosslinked to generate the hydrogel. Cancer cells formed 3D spheroidal structures that underwent near complete dormancy on Amikagel high-throughput drug discovery platform. Due to their dormant status, conventional anticancer drugs such as mitoxantrone and docetaxel that target the actively dividing tumor phenotype were found to be ineffective. Hypothesis driven rational drug discovery approaches were used to identify novel pathways that could sensitize dormant cancer cells to death. Strategies were used to further accelerate the dormant cancer cell death to save time required for the therapeutic outcome.
Amikagel’s properties were chemo-mechanically tunable and directly impacted the outcome of tumor dormancy or relapse. Exposure of dormant spheroids to weakly stiff and adhesive formulation of Amikagel resulted in significant relapse, mimicking the response to changes in extracellular matrix around dormant tumors. Relapsed cells showed significant differences in their metastatic potential compared to the cells that remained dormant after the induction of relapse. Further, the dissertation discusses the use of Amikagels as novel pDNA binding resins in microbead and monolithic formats for potential use in chromatographic purifications. High abundance of amino groups allowed their utilization as novel anion-exchange pDNA binding resins. This dissertation discusses Amikagel formulations for pDNA binding, metastatic cancer cell separation and novel drug discovery against tumor dormancy and relapse.
Amikagel’s properties were chemo-mechanically tunable and directly impacted the outcome of tumor dormancy or relapse. Exposure of dormant spheroids to weakly stiff and adhesive formulation of Amikagel resulted in significant relapse, mimicking the response to changes in extracellular matrix around dormant tumors. Relapsed cells showed significant differences in their metastatic potential compared to the cells that remained dormant after the induction of relapse. Further, the dissertation discusses the use of Amikagels as novel pDNA binding resins in microbead and monolithic formats for potential use in chromatographic purifications. High abundance of amino groups allowed their utilization as novel anion-exchange pDNA binding resins. This dissertation discusses Amikagel formulations for pDNA binding, metastatic cancer cell separation and novel drug discovery against tumor dormancy and relapse.
ContributorsGrandhi, Taraka Sai Pavan (Author) / Rege, Kaushal (Thesis advisor) / Meldrum, Deirdre R (Thesis advisor) / Stabenfeldt, Sarah (Committee member) / Caplan, Michael (Committee member) / Tian, Yanqing (Committee member) / Arizona State University (Publisher)
Created2016
Description
The glucose metabolism level reflects cell proliferative status. A polymeric glucose ratiometric sensor comprising poly(N-(2-hydroxypropyl)methacrylamide) (PHPMA) and poly[2-(methacryloyloxy)ethyl]trimethylammonium chloride (PMAETMA) was synthesized. Cellular internalization and glucose response of the polymer within HeLa cells were investigated.
ContributorsZhang, Liqiang (Author) / Su, Fengyu (Author) / Buizer, Sean (Author) / Kong, Xiangxing (Author) / Lee, Fred (Author) / Day, Kevin (Author) / Tian, Yanqing (Author) / Meldrum, Deirdre (Author) / Biodesign Institute (Contributor)
Created2014-05-07