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Swabs are used as a collecting device for many biological samples and its complete elution is a desired step for clinical and forensic diagnostics. Swabs are made of cotton, rayon, polyurethane, foam or polyester and come in a spun or flocked-tipped format. They are used to extract biological samples from a patient, which includes saliva, mucous, blood, semen or other body fluids. These body fluids then undergo the process of elution where the collected samples are extracted from the swabs into an elution fluid. Apart from biological samples, the importance of swabbing and elution also becomes more evident in forensics, where the concentration of available cells is very low. One such example is rape kit analysis. Another field of application is the capture and release of bacterial spores from environmental contact surfaces and food surfaces, which also indicate the use of swabs in non-biological areas. The recovery of the biological material from the fibre matrix of the swab has a significant influence on diagnostic sensitivity of any assay. The recovery of micro-organisms from a matrix of swab fibres depends on the nature of the body fluid, the type of the swab fibres and the process of elution. Various methods are used to elute samples from swab, including the use of chemicals to digest the cotton fibres to remove intact cells (~20% recovery), centrifugation (~58% recovery), piezoelectric vibration or pressurized fluid-flow (~60% recovery). These methods are either passive (chemical elution) or provides a gentle tangential shear force through associated flow (centrifugation, piezoelectric and pressurized flow), resulting in a low recovery. The success of all the downstream processes of elution, like lysis, DNA amplification and detection, depends on the number of cells eluted from the swab fibre matrix. Hence, the recovery efficiency is an important parameter for determining the performance of elution, and higher value of the same is desired for most diagnostic assays. This thesis reports a magnetically-actuated physical impingement method for elution and recovery of artificial sputum samples from cotton fibres. A device has been fabricated to induce a rotating magnetic field on smaller magnetic particles in a vial for striking the swab within a confined gap. Elution of samples from the swab using this device was demonstrated using artificial sputum prepared by mixing 2% methyl cellulose in deionised water, loaded with fluorescent-tagged polystyrene beads and Escherichia coli bacteria at various concentrations. The recovery efficiency was found to increase with both rotational speed and elution time, but plateaus after 400 RPM and 120s respectively. At higher concentration of polystyrene beads, a maximum recovery of ~85% was achieved at 5x108 particles/ml sample. With lower concentration (106 particles/ml), the maximum efficiency (~93%) was found to be almost twice of the static condition (46.7%), while using only 620µL of elution volume. Similar trends were found in experiments with artificial sputum loaded with E. coli cells, and the maximum recovery was found to be ~90% at 105 CFU/ml concentration. The robust design and smaller size allows the device to be used in different clinical, forensic and laboratory settings. Also, due to cheaper means of manufacturing and assembly, the vials and smaller magnets can be discarded after every experiment, thereby preventing contamination. The device is most suitable for recovering cells from different body fluids like saliva, mucous, semen or blood, absorbed by the swab fibres. Apart from body fluids samples, swabs holding biological agents from environmental surfaces can also be eluted. A higher recovery at lower concentration facilitates the use of this device where the available analyte concentration is low. / Thesis / Master of Applied Science (MASc)

The introduction of ionization under ambient conditions has greatly simplified mass spectrometric analysis. Over past decade, ambient ionization mass spectrometry (MS) methods have revolutionized the way complex samples are analyzed under environmental conditions without requiring, in most cases, any sample pretreatment. Ambient ionization MS gained popularity among other analytical techniques due to its simplicity and its suitability for analysis of small and large molecules. However, ambient ionization methods can suffer from low accuracy and sensitivity due to matrix effects and interferences within complex samples, as well as from poor ionization efficiency. Matrix effects in ambient ionization are usually caused by ion suppression and may depend on different factors, e.g. matrix-to-analyte concentration ratios, proton affinities of analyte and matrix species. To overcome these challenges, in this thesis we present a new approach where a probe is used both as a direct sampling device and as an efficient ambient ionization source. This approach leverages high surface area gold electrodes, fabricated through low-cost bench-top fabrication methods and functionalized using self-assembled alkyl thiol monolayers, as functional conductive sampling probes (FCSPs) for the extraction and concentration of analytes from a sample solution. FCSPs loaded with the targeted analytes were then used to demonstrate a new and highly efficient ionization approach, called Primary Ion Mass Spectrometry Source (PIMSS). In this approach, following capture, the bound analytes are directly desorbed into the mass spectrometer, where ionization is achieved solely through the extraction voltage applied to the probe. 3D-printing was used to design an interface to couple FCSPs to the mass spectrometer. In this work, we discuss a detailed method development and optimization stage and present capabilities of the proposed assay. / Thesis / Master of Science (MSc)

DNA chip technology has drawn tremendous attention since it emerged in the mid 90 s as a method that expedites gene sequencing by over 100-fold. DNA chip, also called DNA microarray, is a combinatorial technology in which different single-stranded DNA (ssDNA) molecules of known sequences are immobilized at specific spots. The immobilized ssDNA strands are called probes. In application, the chip is exposed to a solution containing ssDNA of unknown sequence, called targets, which are labeled with fluorescent dyes. Due to specific molecular recognition among the base pairs in the DNA, the binding or hybridization occurs only when the probe and target sequences are complementary. The nucleotide sequence of the target is determined by imaging the fluorescence from the spots. The uncertainty of background in signal detection and statistical error in data analysis, primarily due to the error in the DNA amplification process and statistical distribution of the tags in the target DNA, have become the fundamental barriers in bringing the technology into application for clinical diagnostics. Furthermore, the dye and tagging process are expensive, making the cost of DNA chips inhibitive for clinical testing. These limitations and challenges make it difficult to implement DNA chip methods as a diagnostic tool in a pathology laboratory. The objective of this dissertation research is to provide an alternative approach that will address the above challenges.. In this research, a label-free assay is designed and studied. Polystyrene (PS), a commonly used polymeric material, serves as the fluorescence agent. Probe ssDNA is covalently immobilized on polystyrene thin film that is supported by a reflecting substrate. When this chip is exposed to excitation light, fluorescence light intensity from PS is detected as the signal. Since the optical constants and conformations of ssDNA and dsDNA (double stranded DNA) are different, the measured fluorescence from PS changes for the same intensity of excitation light.. The fluorescence contrast is used to quantify the amount of probe-target hybridization. A mathematical model that considers multiple reflections and scattering is developed to explain the mechanism of the fluorescence contrast which depends on the thickness of the PS film. Scattering is the dominant factor that contributes to the contrast. The potential of this assay to detect single nucleotide polymorphism is also tested. / Ph. D.

A numerical analysis based on the finite element method is developed to simulate the mechanical response of a typical sixteen-ounce plastic drink cup subjected to a lateral compressive load. The aim of the analysis is to simulate a test in which the cup is supported horizontally in a fixture on a testing machine platen, and a loading nose attached to the actuator is displaced downward into the cup. The numerical model is developed using the software packages MSC.Patran, ABAQUS/CAE, and ABAQUS/Standard. The high impact polystyrene material of the cup is modeled as linear elastic, considering isotropic and orthotropic material behavior. The structural model of the cup is a truncated conical shell including a ring at the open end of the cup and circumferential stiffening ribs. The analysis is based on small strain, large rotation shell kinematics, and the loading apparatus of the test is simulated with a rigid, circular cylinder contacting the cup. Coupons cut from the wall of a cup are subjected to tension to determine the ranges of the meridional and circumferential moduli of elasticity. Rings cut from the open end of the cup were tested in diametrical tension to aid in validating the finite element modeling. Reasonable correlation of the simulation to available cup compression test data is achieved. Parametric studies are conducted for several meridional thickness distributions of the cup wall, and for a range of orthotropic material properties. / Master of Science