<p> The goal of this research project was the development of a protocol for preparation of optically clear organic/inorganic hybrid materials that was amenable to entrapment of lipophilic biomolecules. The protocol involved the acid-catalyzed hydrolysis of mixtures of tetraethylorthosilicate (TEOS) with organosilane precursors, including methyltriethoxysilane (MTES), dimethyldimethoxysilane (DMDMS) and propyltrimethoxysilane (PTMS) in the presence and absence of the polymer additives poly(ethylene glycol) or poly(vinyl alcohol).</p> <p> The effect of organosilane precursors and polymer additives on the optical clarity, hardness and hydration stability of the resulting materials was characterized. It was determined that there was a limit to the amount of organosilane that could be added before the materials exhibited unacceptable characteristics. These limits were 20.0% (v/v) for MTES, 10.0% (v/v) for PTMS, and 5.0% (v/v) for DMDMS. Addition of PEG to these materials at levels up to 10.0% (w/v) resulted in good material characteristics. However, addition of PVA produced opaque materials with poor material properties. The internal environment of the materials was also probed using the environmentally sensitive fluorescent probes 7-azaindole (7AI) and prodan. These studies showed that the method of hydrolysis of the silane precursors and the aging conditions had a dramatic effect on the resulting material.</p> <p> The hybrid materials were used to entrap human serum albumin (HSA) and lipase to determine the effect of organic content on the biological function of these biomolecules. Both biomolecules retained a portion of their native function when entrapped in sol-gel-derived materials, and it was found that both proteins showed enhanced function in the presence of MTES. In the case of lipase, it was also determined that addition of PEG 600 at 10.0% (w/v in the gelation buffer) provided a dramatic increase in activity compared to materials without this additive, likely owing to a direct effect of the PEG on the stability of the entrapped protein.</p> <p> Following studies using bulk glasses, a protocol was developed for the preparation of optically clear sol-gel-derived thin films that was amenable to entrapment of biomolecules. The optimal method involved dipcasting of co-hydrolyzed materials containing 1.0 to 3.0% PEG. By careful control of the viscosity of the casting solution and the rate of film deposition, it was possible to form very stable thin films with excellent physical characteristics. These films were used to entrap the pH-sensitive, ratiometric fluorescent probe dextran-SNARF-1, resulting in a prototype of a fluorimetric pH sensor. Co-entrapment of the probe and lipase into sol-gel-derived thin films resulted in a rapid, reagentless biosensor prototype that could monitor changes in pH due to the enzyme-catalyzed hydrolysis of triglycerides. These results demonstrate that species entrapped in sol-gel derived thin films are suitable for biosensor development.</p> / Thesis / Master of Science (MSc)

Conventional methods for diagnosis of human disease are, at times, limited in different regards including time requirement, either experimental or data processing, sensitivity, and selectivity. It is then that a Point of Care Criteria, which considers the true utility and usefulness of the device, is employed to propose new diagnostic devices capable of overcoming the aforementioned shortcomings of conventional tools. Nucleic acid, characterized for its predictable base-pairing nature, is considered to be a highly-selective, yet greatly modifiable device. Its behavior is then described through Boolean Logic, where “true” or “false” outputs are mathematically described as “1” and “0”, respectively. This mathematical approach is then referred to as Logic Gates, where outputs can be predicted based on satisfied environmental conditions. The mechanisms, capable of exhibiting Logic Gate behavior, are described.

No description available.

No description available.

Raman spectroscopy and its counterpart surface-enhanced Raman scattering (SERS) have proven to be effective methods for detecting miniscule changes in the phenotypes of E. coli and other single-celled organisms to aid in the detection of new strains for industrial use and discovery of new antibiotics. The purpose of this study is to develop a method to quickly and accurately detect contaminants in water samples through phenotype changes in E. coli measured through SERS. Contaminated Luria-Bertani (LB) media was inoculated with LB with an OD600 of 1, grown for two hours, and then dried on a flat piece of aluminum foil. These samples were then Raman scanned and processed to determine contaminant-induced changes to the phenotypes of the E. coli. Three types of tests were run to show the effectiveness of this method: single-component, multicomponent, and impure water sources. In single-component tests, it was found that differences due to NaCl contamination could be detected to 5.0E-9 weight percent (wt %), ethanol (EtOH) to 5.0E-7 volumetric percent (% v/v), citric acid (CA) to 2.8E-4 wt %, acetic acid (AA) to 2.6E-4 wt %, kanamycin to 2.5E-11 wt %, ampicillin to 2.5E-10 wt %, CoCl2 to trace amounts, and silver nanoparticles (AgNP) to 5.2E-7 wt %. Many of these are below the detection limits of analytical instrumentation, but their effects on E. coli phenotypes were detectable by Raman spectroscopy. Multicomponent tests showed that in a mixture, the most toxic or most concentrated contaminants have the most effect on cell phenotype. However, it was shown that similar concentrations of similar contaminants may be difficult to discern with current methods. This behavior was also seen in the impure water samples, showing that tap water behaves the closest to a DI control, followed by running water, and finally stagnant bodies. This new method of monitoring E. coli phenotypes with Raman spectroscopy as a biosensor shows promise for the fast, portable, and accurate determination of environmental contaminants with a broad-spectrum and very low detection limits. / Master of Science / Recently, Raman spectroscopy and an enhanced version called surface-enhanced Raman scattering (SERS) have shown promise in showing the effects of a cell’s environment on how it expresses genes and what chemical compounds it produces to survive. It does this through reading the chemical signature it gives off due to these changes, and these readings have been used for showing the effects of antibiotics, finding varieties that are resistant to toxic byproducts of their activities, and as biosensors. The study outlined in this thesis aims to develop a test utilizing SERS to see the reactions of a non-pathogenic strain of E. coli to contaminants in their media and determine their identity. This test was run for three types of contaminated samples: one contaminant, three contaminants, and tests using impure water from a sink tap, a pond, and a stream. For the single contaminants, eight types were tested; NaCl, ethanol, citric acid, acetic acid (AA), kanamycin, ampicillin, CoCl2, and silver nanoparticles. Detection limits for all contaminants were found, with the lowest detectable concentrations all falling below or matching detection limits of common methods. The lowest detectable concentrations came from kanamycin and CoCl2, at 2.5E-11 weight percent and in amounts which are considered beneficial to the environment, respectively. The three-contaminant test shows that it is possible to pinpoint which contaminants are having the highest effect, though if contaminants are similar in nature and in similar concentrations, it may be difficult to pinpoint which is causing the effect. In the final test, similarity of water sources to pure water were found, with tap water being closest, followed by stream water and the most different being pond water. It was also found that pond water and stream water were closest in behavior, showing the power of this method in differentiating sources from each other.

Parametric design models were created to optimize cable sensitivities in a structural health-monitoring system. Experiments were also conducted to determine the sensitivity of a working system. The system consists of time domain reflectometry (TDR) signal processing equipment and specially designed sensing cables. The TDR equipment sends a high-frequency electric pulse (in the gigahertz range) along the sensing cable. Any change in electric impedance along the cable reflects some portion of the electric pulse back to the TDR equipment. The time delay, amplitude, and shape of the reflected pulse provides the means to respectively locate, determine the magnitude, and indicate the nature of the change in impedance. The change in the sensing cable impedance may be caused by cable elongation (change in resistance), shear deformation (change in capacitance), corrosion of the cable or the materials around the cable (change in inductance), etc. The sensing cables are an essential part of the health-monitoring system because the cable design parameters determine the cable impedance sensitivity to structural changes. By using parametric design models, the optimum cable parameters can be determined for specific cases and used to select or design an appropriate cable. Proof-of-concept and resolution experiments were also conducted to provide, respectively, verification and sensitivity of the system. / Master of Science

No description available.

Neurodegenerative disease (ND) is a collection of progressive disorders which is marked by gradual degeneration of the central nervous system (CNS). The damage of brain and nervous system causes diminishing of brain and nerve cell which lead to body dysfunction, organ failure, paralysis and ultimately death of a patient. Neurodegenerative diseases affect millions of people worldwide. United States of America is among highest ND prevailing disease countries. Yet the number of cases is underreported because of the complexity of disease diagnosis. With the increase in aging population the rate of ND cases is also increasing. Another important concern is that viral pandemic like COVID-19 may additionally contribute to rise in ND affected population. Currently there is no cure available for ND except a few treatments which decrease some symptoms and decrease disease progression. Early-stage diagnosis of ND is therefore an important and immediate area of research which can improve the quality of life of affected people and help in health management. Many ND has been associated with abnormal expansion of tandem repeat sequence of nucleic acid and other associated protein biomarkers. The discovery of these biomarkers holds promise for improving diagnosis, treatment, and efficacy monitoring. Current detection methods such as neuroimaging techniques, Real-Time quantitative Polymerase Chain Reaction (RT-qPCR) and Enzyme-Linked Immunosorbent Assay (ELISA) are complex, expensive, require laboratory setup and/or trained operator. Electrochemical biosensing offers a promising portable, inexpensive, and sensitive platform for early and robust diagnosis and healthcare management. In this dissertation a label free electrochemical method is developed to investigate the effect of complex targets for potential genosensing applications and effect of complex biomatrix for immunosensing applications. We first prepare sensing surfaces with three different backbone microprobe nucleic acids to detect length- and sequence-dependent complex secondary structures containing RNA linked to Huntington’s disease, based on the charge transfer resistance of the interface. Then we reported an immunosensing surface using commercially available screen-printed electrode to detect ALS associated biomarker neurofilament light chain in serum samples. Finally, we establish an in-house and cost effective electrochemical immunosensing platform for the detection of ALS biomarker poly-glycine-proline dipeptide repeat in cerebrospinal fluid.

ABSTRACT BIOACTIVE SURFACE DESIGN BASED ON CONDUCTING POLYMERS AND APPLICATIONS TO BIOSENSORS Ekiz, Fulya M. Sc., Department of Biotechnology Supervisor: Prof. Dr. Levent Toppare Co-Supervisor: Prof. Dr. Suna Timur June 2012, 88 pages An underlying idea of joining the recognition features of biological macromolecules to the sensitivity of electrochemical devices has brought the concept of biosensors as remarkable analytical tools for monitoring desired analytes in different technological areas. Over other methods, biosensors have some advantages including high selectivity, sensitivity, simplicity and this leads to solutions for some problems met in the measurement of some analytes. In this context, conducting polymers are excellent alternatives with their biocompatibility and ease of applicability for an efficient immobilization of biomolecules in preparing biosensors. Using several materials and arranging the surface properties of the electrodes, more efficient and seminal designs can be achieved. In this thesis, it is aimed to create new direct biosensors systems for the detection of several analytes such as glucose and pesticides thought to be harmful to the environment. Recently synthesized conducting polymers (polyTBT) / (poly(2-dodecyl-4,7-di(thiophen-2-yl)-2H-benzo[ d][1,2,3]triazole) and (poly(TBT 6 -NH2 ) / poly(6-(4,7-di(thiophen-2-yl)-2H-benzo[d][1,2,3]triazol-2-yl)hexan-1-amine) were utilized as a matrices for biomolecule immobilization. After successful electrochemical deposition the polymers on the graphite electrode surfaces, immobilization of glucose oxidase (GOx) and choline oxidase (ChO) were carried out. Amperometric measurements were recorded by monitoring oxygen consumption in the presence of substrates at -0.7 V. The optimized biosensors showed a very good linearity with rapid response times and low detection limits (LOD) to glucose and choline. Also, kinetic parameters, operational and storage stabilities were determined. Finally, designed biosensor systems were applied for glucose and pesticide detection in different media.

<p>The development of low-cost, and reliable platforms for on-site detection of pathogenic agents, and toxic environmental traces is still a critical need for real-time monitoring of potential environmental pollution and imminent outbreaks. The biosensors market is projected to attain 31.5 billion by 2024. In this landscape, colorimetric and electrochemical devices continue to have significant relevance, with paper-based platforms leading the point-of-care (POC) segment for pathogen detection and environmental monitoring.</p> <p>Despite the true potential of biosensors in general, they have witnessed a slow rate in commercialization, mainly due to cost restrictions, and concerns related to their reliability and repeatability once scaled-up. This research evaluates the implementation of printing techniques as a strong approach for the fabrication of paper-based and flexible electrochemical biosensors. The results obtained demonstrated the ability to control and predict the variables affecting the sensing performance, achieving high precision of the printing parameters, and allowing optimization, and iterations since very early stages of prototype development.</p> <p>Besides the novel fabrication approach, this work introduces the use of truncated aptameric DNA sequences for whole cell detection of E. coli O157:H7 and heavy metals (Hg2+ and As3+), providing evidence of high stability and robustness under harsh conditions. Results obtained demonstrate their equal or even superior performance when compared to antibodies.</p> <p>We established the use of aptamer-functionalized multilayered label particles (PEI-grafted gold decorated polystyrene) with high stability as label particles. These particles address the well known drawback of non-selective aggregation typical of traditional naked Gold nanoparticles. The outstanding stability of these multilayered labels was demonstrated when used in an enhanced version of the lateral flow assay for detection of E. coli O157:H7 (state of the art for paper-based colorimetric detection of whole cell bacteria), and in a multiplexed paper-based microfluidic device for dual detection of Mercury and Arsenic. This work sets the foundation of the development of a next generation of health care and environmental monitoring devices that are portable, sensitive, quantitative, and can reliably detect multiple targets with one single test.</p>