Key Words: Thrombosis, Vascular Graft, Microfluidics, Wall Shear Stress / Vascular grafts are essential for the management of cardiovascular disease. However, the lifesaving potential of these devices is undermined by thrombosis arising from material and flow interactions on the blood contacting surface. To combat this issue, the use of antithrombogenic coatings has emerged as a promising strategy for modulating blood and graft interaction in vivo. Although an important determinant of graft performance, hemodynamics are frequently overlooked in the in vitro testing of coatings and their translatability remains poorly understood. We address this limitation with a microscale platform that incorporates vascular prosthesis and coatings with tuneable flow and surface conditions in vitro. As a proof of concept, we use the platform to test the thrombogenic performance of a novel class of lubricant infused (LIS) and antibody lubricant infused (anti-CD34 LIS) coated ePTFE vascular grafts in the presence of arterial wall shear stress, with and without the presence of endothelial cells. Our findings suggest lubricant infused coated ePTFE vascular grafts are thromboresistant under flow and may have potential for in vivo arterial grafting applications. It is moreover apparent that the microscale properties of the device could be advantageous for the testing and translation of novel antithrombogenic coatings or blood contacting prosthesis in general. / Thesis / Master of Applied Science (MASc)

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Magnetic particles with active functional groups offer numerous advantages for use in μ-TAS (Micro Total Analytical Systems). The functional site allows chemical binding of the particle with the target species in the fluid sample. Selection of the functional group establishes the target molecule and vice versa under assumptions of highly specific biding. The particles hence act as mobile reaction substrates with high surface to volume ratios owing to their small size. The concept of action at a distance allows their use as agents for separation in microchannels based on relatively simple design. It is possible to manipulate magnetic particles and bound target species using an externally applied magnetic field. Hence, the particles can be effectively separated from the flow of a carrier fluid. Magnetic fields create dipolar interactions causing the particles to form interesting structures and aggregates. Depending upon the applied field, the microstructure evolution of the aggregate is interesting in its own right, e.g. related to improvements in material properties and bottom-up self assembly. The shape of the aggregates can be determined a priori if the interaction between the particles is well characterized. The dominant competing forces that influence magnetic particle dynamics in a flow are magnetic and viscous. There are a number of physical parameters such as viscosity, magnetic susceptibility, fluid velocity, etc. which are varied to study their individual effects. Initially dilute suspensions are studied experimentally and numerically using a particle based dynamics approach. Once established, a force model for particle interaction is investigated for concentrated suspensions. A Lagrangian particle tracking algorithm that returns positions of the particles is used for this work that focuses on studying the dynamics of these particles. A mathematical model is proposed and investigated for functionalization between magnetic and non-magnetic particles. Having characterized the collection of magnetic particles, the effect of relative concentrations is investigated on the collection of the non-magnetic species. / Ph. D.

Mortality due to cancer is a global health issue that can be improved through further development of diagnostic and prognostic tools. Recent advancements in technologies aiding cancer research have made significant strides, however a demand for a non-invasive clinically relevant point-of-care tools exists. To accomplish this feat, the desired instrument needs to be low-cost, easy-to-operate, efficient, and have rapid processing and analysis. Microfluidic platforms in cancer research have proven to be advantageous due to its operation at the microscale, which has low costs, favorable physics, high precision, short experimentation time, and requires minimal reagent and sample sizes. Label-free technologies rely on cell biophysical characteristics to identify, evaluate, and study biological samples. Biomechanical probing of cells through deformability assays provides a label-free method of identifying cell health and monitoring response to physical and chemical stimuli. Bioimpedance analysis is an alternative versatile label-free method of evaluating cell characteristics by measuring cell response to electrical signals. Microfluidic technologies can facilitate biomechanical and bioelectrical analysis through deformability assays and impedance spectroscopy. This dissertation demonstrates scientific contributions towards single-cell analysis and liquid biopsy devices focusing on cancer research. First, cell deformability assays were improved through the introduction of multi-constriction channels, which revealed that cells have a non-linear response to deformation. Combining impedance analysis with microfluidic deformability assays provided a large dataset of mechano-electrical information, which improved cell characterization and greatly decreased post-processing times. Next, two unique biosensors demonstrated improved throughput while maintaining sensitivity of single-cell analysis assays through parallelization and incorporating machine learning for data processing. Liquid biopsies involve studying cancer cells in patient vascular systems, called circulating tumor cells (CTCs), through blood samples. CTC tests reveal valuable information on patient prognosis, diagnosis and can aide therapy selection in a minimally invasive manner. This body of work presents two liquid biopsy devices that enrich murine and human blood samples and isolate CTCs to ease detection and analysis. Additionally, a microfluidic CTC detection biosensor is introduced to reliably count and identify cancer cells in murine blood, where an extremely low-cost version of the assay is also validated. Thus, the assays presented in this dissertation show promise of microfluidic technologies towards point-of-care systems for cancer research. / Doctor of Philosophy / Cancer is the second leading cause of death worldwide with approximately 2 million new cases each year in the just United States. Significant research development for diagnostic and prognostic tools have been conducted, however they can be expensive, invasive, time-consuming, unreliable, and not always easily accessible. Thus, a tool that is cheap, minimally invasive, easy-to-use, and robust needs to be developed to combat these issues. Typical cancer studies have primarily focused on biological and biochemical methods for evaluation; however, researchers have begun to leverage small-scale biosensors that utilize biophysical attributes. Recent studies have proven that these lab-on-a-chip technologies can produce meaningful results by exploiting these biophysical characteristics. Microfluidics is a science that consists of sub-millimeter sized channels which show a great deal of promise as they require minimal materials and can quickly and efficiently analyze biological samples. Label-free methods of studying cells rely on their physical properties, such as size, deformability, density, and electrical properties. These biophysical characteristics can be easily obtained at the single-cell level through microfluidic-based assays. Measuring and monitoring these attributes can provide valuable information to help understand cancer cell response to stimuli such as chemotherapeutic drugs or other therapies. A liquid biopsy is a non-invasive method of evaluating cancer patients by studying circulating tumor cells (CTCs) that exist in their blood. This dissertation reports a wide range of label-free microfluidic assays that evaluate and study biological samples at the single-cell level and for liquid biopsies. These assays consist of microfluidic channels with sensors that can rapidly obtain biophysical characteristics and process blood samples for liquid biopsy applications. Uniquely modifying microfluidic channel geometries and sensor configurations improved upon previously developed single-cell and CTC-based tools. The resulting devices were low in cost, easy-to-use, efficient, and reliable methods that alleviates current issues in cancer research while showing clinical utility.

The discovery of cancer biomarkers is crucial in the clinical setting to facilitate early diagnosis and treatment, thereby increasing survival rates. Proteomic technologies with mass spectrometry detection (MS) have the potential to affect the entire spectrum of cancer research by identifying these biomarkers. Simultaneously, microfabricated devices have evolved into ideal analysis platforms for minute amounts of sample, with promising applications for proteomic investigations and future biomarker screening. This thesis reports on the analysis of the proteomic constituents of the MCF7 breast cancer cell line using a shotgun 2-D strong cationic exchange/reversed phase liquid chromatography electrospray ionization tandem mass spectrometry (SCX/RP-LC-ESI-MS/MS) protocol. A series of optimization strategies were performed to improve the LC-MS experimental set-up, sample preparation, data acquisition and database searching parameters, and to enable the detection and confident identification of a large number of proteins. Over ~4,500 proteins were identified using conventional filtering parameters, and >2000 proteins using a combination of filters and p-value sorting. Of these, ~1,950 proteins had p<0.001 (~90%) and more than half were identified by &#8805; 2 unique peptides. About 220 proteins were functionally involved in cancer related cellular processes, and over 100 proteins were previously described in the literature as potential cancer markers. Biomarkers such as PCNA, cathepsin D, E-cadherin, 14-3-3-sigma, antigen Ki-67, TP53RK, and calreticulin were identified. These data were generated by subjecting to mass spectrometric analysis ~42 µg of protein digest, analyzing 16 SCX peptide fractions, and interpreting ~55,000 MS2 spectra. Total MS time required for analysis was 40 h. Selective SCX fractions were also analyzed by using a microfluidic LC platform. The performance of the microchip LC was comparable to that obtained with bench-top instrumentation when similar experimental conditions were used. The identification of 5 cancer biomarkers was enabled by using the microchip LC platform. Furthermore, this device was also capable to analyze phosphopeptides. / Master of Science