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Full-Text Articles in Engineering
Engineering Cell-Free Protein Synthesis Biosensors For Point Of Care Use, Tyler Jordan Free
Engineering Cell-Free Protein Synthesis Biosensors For Point Of Care Use, Tyler Jordan Free
Theses and Dissertations
Diagnostic tests can dramatically improve and save lives. Survival rates and patient outcomes related to cancer and other diseases can be improved when routine testing guides personalized treatment. Highly accurate laboratory tests exist for thousands of biomarkers, including those of interest in this dissertation, but cost and complexity limit access to life-saving lab tests in many cases throughout the world. To fill this need and increase accessibility of diagnostics, in this report we report our engineering advancements towards portable biosensors for colorimetric detection of glutamine, endocrine disrupting compounds (EDC), and pathogenic RNA which are key biomarkers in cancer treatment monitoring, …
Validation And Development Of Top-Down Illumination For Optofluidic Biosensors, Matthew Marley Hamblin
Validation And Development Of Top-Down Illumination For Optofluidic Biosensors, Matthew Marley Hamblin
Theses and Dissertations
Lab-on-a-chip devices are changing the way that medical testing is performed by allowing rapid testing with small samples. Optofluidic biosensors are a type of lab-on-a-chip device that use light excitation on a fluid sample. One such application of an optofluidic biosensor is a device that can detect antibiotic resistant bacteria by combining DNA from a sample with fluorescent beads, flowing that sample through a hollow channel, and shining laser light on the channel. If the bacteria tested for is present, the fluorescent beads will give off photons that can be detected as a positive signal. The main method for illumination …
Optofluidic Manipulation With Nanomembrane Platforms Used For Solid-State Nanopore Integration, Zachary J. Walker
Optofluidic Manipulation With Nanomembrane Platforms Used For Solid-State Nanopore Integration, Zachary J. Walker
Theses and Dissertations
Nanopore technology has introduced new techniques for single particle detection and analysis. A nanopore consists of a small opening in a membrane on the nanometer scale. Nanopores are found in nature and are utilized for transporting molecules through biological membranes. Researchers have been able to mimic naturally forming biological nanopores and utilize them for a variety of sensing applications. Nanopores, fabricated either organically or inorganically, can be used for detecting biomarkers such as proteins, nucleic acids, and metabolites that translocate the membrane by way of the nanopore. Constant ionic current flow is measured through the nanopore by way of a …
An Integrated Model Of Optofluidic Biosensor Function And Performance, Joel Greig Wright, Jr.
An Integrated Model Of Optofluidic Biosensor Function And Performance, Joel Greig Wright, Jr.
Theses and Dissertations
Optofluidic flow-through biosensor devices have been in development for fast bio-target detection. Utilizing the fabrication processes developed by the microelectronics industry, these biosensors can be fabricated into lab-on-a-chip devices with a degree of platform portability. This biosensor technology can be used to detect a variety of targets, and is particularly useful for the detection single molecules and nucleic acid strands. Microfabrication also offers the possibility of production at scale, and this will offer a fast detection method for a range of applications with promising economic viability. The development of this technology has advanced to now warrant a descriptive model that …
Designing Cell-Free Protein Synthesis Systems For Improved Biocatalysis And On-Demand, Cost-Effective Biosensors, Mehran Soltani Najafabadi
Designing Cell-Free Protein Synthesis Systems For Improved Biocatalysis And On-Demand, Cost-Effective Biosensors, Mehran Soltani Najafabadi
Theses and Dissertations
The open nature of Cell-Free Protein Synthesis (CFPS) systems has enabled flexible design, easy manipulation, and novel applications of protein engineering in therapeutic production, biocatalysis, and biosensors. This dissertation reports on three advances in the application of CFPS systems for 1) improving biocatalysis performance in industrial applications by site-specific covalent enzyme immobilization, 2) expressing and optimizing a difficult to express a mammalian protein in bacterial-based CFPS systems and its application for cost-effective, on-demand biosensors compatible with human body fluids, and 3) streamlining the procedure of an E. coli extract with built-in compatibility with human body fluid biosensors. Site-specific covalent immobilization …
Engineering Cell-Free Protein Expression Systems For Biotherapeutics And Biosensing, John Porter Hunt
Engineering Cell-Free Protein Expression Systems For Biotherapeutics And Biosensing, John Porter Hunt
Theses and Dissertations
Therapeutic proteins have become a cornerstone of modern medicine since the FDA approval of recombinant human insulin in 1982. Likewise, biosensors transform chemical detection and disease diagnostics by identifying biomarkers, chemical contaminants, and infective agents. Long-standing methods for creating therapeutics and biosensors employ whole cells such as Escherichia coli (E. coli). Alternatively, cell-free protein synthesis (CFPS) employs the enzymatic reactions necessary for protein production and biosensing within a cell, but in an engineered reactor environment facilitating unprecedented access to and control over biochemical machinery, preservation by cryodesiccation for portable deployment, and functionality in cytotoxic applications. This dissertation reports advances in …
3d Interdigitated Vertically Aligned Carbon Nanotube Electrodes For Electrochemical Impedimetric Biosensing, Benjamin J. Brownlee, Jonathan C. Claussen, Brian D. Iverson
3d Interdigitated Vertically Aligned Carbon Nanotube Electrodes For Electrochemical Impedimetric Biosensing, Benjamin J. Brownlee, Jonathan C. Claussen, Brian D. Iverson
Faculty Publications
Advances in nanomaterials, combined with electrochemical impedance spectroscopy (EIS), have allowed electrochemical biosensors to have high sensitivity while remaining labe-lfree, enabling the potential for portable diagnosis at the point-of-care. We report porous, 3D vertically aligned carbon nanotube (VACNT) electrodes with underlying chromium electrical leads for impedance-based biosensing. The electrodes are characterized by electrode height (5, 25, and 80 μm), gap width (15 and 25 μm), and geometry (interdigitated and serpentine) using scanning electron microscopy, cyclic voltammetry, and EIS. The protein streptavidin is functionalized onto VACNT electrodes for detection of biotin, as confirmed by fluorescence microscopy. EIS is used to measure …
Three-Dimensional Hydrodynamic Focusing For Integrated Optofluidic Detection Enhancement, Erik Scott Hamilton
Three-Dimensional Hydrodynamic Focusing For Integrated Optofluidic Detection Enhancement, Erik Scott Hamilton
Theses and Dissertations
The rise of superbugs, including antibiotic-resistant bacteria, and virus outbreaks, such as the recent coronavirus scare, illustrate the need for rapid detection of disease pathogens. Widespread availability of rapid disease identification would facilitate outbreak prevention and specific treatment. The ARROW biosensor microchip can directly detect single molecules through fluorescence-based optofluidic interrogation. The nature of the microfluidic channels found on optofluidic sensor platforms sets some of the ultimate sensitivity and accuracy limits and can result in false negative test results. Yet higher sensitivity and specificity is desired through hydrodynamic focusing. Novel 3D hydrodynamic focusing designs were developed and implemented on the …
Multiplexed Optofluidics For Single-Molecule Analysis, Matthew Alan Stott
Multiplexed Optofluidics For Single-Molecule Analysis, Matthew Alan Stott
Theses and Dissertations
The rapid development of optofluidics, the combination of microfluidics and integrated optics, since its formal conception in the early 2000's has aided in the advance of single-molecule analysis. The optofluidic platform discussed in this dissertation is called the liquid core anti-resonant reflecting optical waveguide (LC-ARROW). This platform uses ARROW waveguides to orthogonally intersect a liquid core waveguide with solid core rib waveguides for the excitation of specifically labeled molecules and collection of fluorescence signal. Since conception, the LC-ARROW platform has demonstrated its effectiveness as a lab-on-a-chip fluorescence biosensor. However, until the addition of optical multiplexing excitation waveguides, the platform lacked …
Improved Single Molecule Detection Platform Using A Buried Arrow Design, Thomas Allen Wall
Improved Single Molecule Detection Platform Using A Buried Arrow Design, Thomas Allen Wall
Theses and Dissertations
As the microelectronics industry pushes microfabrication processes further, the lab-on-a-chip field has continued to piggy-back off the industry's fabrication capabilities with the goal of producing total chemical and biological systems on small chip-size platforms. One important function of such systems is the ability to perform single molecule detection. There are currently many methods being researched for performing single molecule detection, both macro and micro in scale. This dissertation focuses on an optofluidic, lab-on-a-chip platform called the ARROW biosensor, which possesses several advantages over macro-scale single molecule detection platforms. These advantages include an amplification-free detection scheme, cheap parallel fabrication techniques, rapid …
Engineering Cell-Free Protein Synthesis Technology For Codon Reassignment, Biotherapeutics Production Using Just-Add-Water System, And Biosensing Endocrine Disrupting Compounds, Sayed Mohammad Salehi
Engineering Cell-Free Protein Synthesis Technology For Codon Reassignment, Biotherapeutics Production Using Just-Add-Water System, And Biosensing Endocrine Disrupting Compounds, Sayed Mohammad Salehi
Theses and Dissertations
Cell-free protein synthesis is an emerging technology that has many applications. The open nature of this system makes it a compelling technology that can be manipulated to answer many needs that are unavailable in other systems. This dissertation reports on engineering this technology for: 1) sense codon emancipation for incorporation of multiple unnatural amino acids; 2) expressing a hard-to-express anticancer biotherapeutic and introducing a just-add-water system; 3) a biosensing ligand that interacts with nuclear hormone receptors. Emancipating sense codons toward a minimized genetic code is of significant interest to science and engineering. A promising approach to sense codon emancipation is …
Microfluidic Devices And Biosensors, Long-Fang Tsai
Microfluidic Devices And Biosensors, Long-Fang Tsai
Theses and Dissertations
My research broadly covers various important aspects of microfluidic devices and biosensors. Specifically, this dissertation reports: (1) a new and effective room temperature method of bonding polydimethylsiloxane (PDMS) microfluidics to substrates such as silicon and glass, (2) a new microfluidic pump concept and implementation specifically designed to repeatedly drive a small sample volume (<1 µL) very rapidly (~500 µL/min) through a sensor-containing flow channel to significantly decrease sensor response time through advection-driven rather than diffusion-driven mass transport, (3) use of a new microfluidic material based on polyethylene glycol diacrylate (PEGDA) to implement impedance-based dynamic nanochannel sensors for protein sensing, and (4) an investigation of galvanoluminescence and how to avoid it for conditions important to fluorescence-based dielectrophoresis (DEP) microfluidic biosensors. Over the last decade, the Nordin research group has developed a lab-on-a-chip (LOC) biosensor based on silicon photonic microcantilever arrays integrated with polydimethylsiloxane (PDMS) microfluidics for protein biomarker detection. Integration requires reliable bonding at room temperature with adequate bond strength between the PDMS element and microcantilever sensor substrate. The requirement for a room temperature process is particularly critical because microcantilevers must be individually functionalized with antibody-based receptor molecules prior to bonding and cannot withstand significant heating after functionalization. I developed a new room temperature bonding method using PDMS curing agent as an intermediate adhesive layer. Two curing agents (Sylgard 184 and 182) were compared, as well as an alternate UV curable adhesive (NOA 75). The bond strength of Sylgard 184 was found to be stronger than Sylgard 182 under the same curing conditions. Overnight room temperature curing with Sylgard 184 yields an average burst pressure of 433 kPa, which is more than adequate for many PDMS sensor devices. In contrast, UV curable epoxy required a 12 hour bake at 50 °C to achieve maximum bond strength, which resulted in a burst pressure of only 124 kPa. In many biosensing scenarios it is desirable to use a small sample volume (<1 µL) to detect small analyte concentrations in as short a time as possible. I report a new microfluidic pump to address this need, which we call a reflow pump. It is designed to rapidly pump a small sample volume back and forth in a flow channel. Ultimately, the flow channel would contain functionalized sensor surfaces. The rapid flow permits use of advection-driven mass transport to the sensor surfaces to dramatically reduce sensor response times compared to diffusion-based mass transport. Normally such rapid flow would have the effect of decreasing the fraction of analyte molecules in the volume that would see the sensor surfaces. By configuring the pump to reflow fluid back and forth in the flow channel, the analyte molecules in the small sample volume are used efficiently in that they have many opportunities to make it to the sensor surfaces. I describe a 3-layer PDMS reflow pump that pumps 300 nL of fluid at 500 µL/min for 15 psi actuation pressure, and demonstrate a new two-layer configuration that significantly simplifies pump fabrication. Impedance-based nanochannel sensors operate on the basis of capturing target molecules in nanochannels such that impedance through the nanochannels is increased. While simple in concept, the response time can be quite long (8~12 hours) because the achievable flow rate through a nanochannel is very limited. An approach to dramatically increase the flow rate is to form nanochannels only during impedance measurements, and otherwise have an array of nanotrenches on the surface of a conventional microfluidic flow channel where they are exposed to normal microfluidic flow rates. I have implemented such a dynamic nanochannel approach with a recently-developed microfluidic material based polyethylene glycol diacrylate (PEGDA). I present the design, fabrication, and testing of PEGDA dynamic nanochannel array sensors, and demonstrate an 11.2 % increase in nanochannel impedance when exposed to 7.2 µM bovine serum albumin (BSA) in phosphate buffered saline (PBS). Recently, LOC biosensors for cancer cell detection have been demonstrated based on a combination of dielectrophoresis (DEP) and fluorescence detection. For fluorescence detection it is critical to minimize other sources of light in the system. However, reported devices use a non-noble metal electrode, indium tin oxide (ITO), to take advantage of its optical transparency. Unfortunately, use of non-noble metal electrodes can result in galvanoluminescence (GL) in which the AC voltage applied to the electrodes to achieve DEP causes light emission, which can potentially confound the fluorescence measurement. I designed and fabricated two types of devices to examine and identify conditions that lead to GL. Based on my observations, I have developed a method to avoid GL that involves measuring the impedance spectrum of a DEP device and choosing an operating frequency in the resistive portion of the spectrum. I also measure the emission spectrum of twelve salt solutions, all of which exhibited broadband GL. Finally, I show that in addition to Au, Cr and Ni do not exhibit GL, are therefore potentially attractive as low cost DEP electrode materials.
Functionalization Of In-Plane Photonic Microcantilever Arrays For Biosensing Applications, Stanley J. Ness
Functionalization Of In-Plane Photonic Microcantilever Arrays For Biosensing Applications, Stanley J. Ness
Theses and Dissertations
Microcantilevers have been investigated as high sensitivity, label free biosensors for approximately 15 years. In nearly all cases, a thin gold film deposited on the microcantilevers is used as an intermediate attachment layer because of the convenience of thiol-gold chemistry. Unfortunately, this attachment chemistry can be unstable when used with complex sample media such as blood plasma. The Nordin group at BYU has recently developed an all-silicon in-plane photonic microcantilever (PMCL) technology to serve as a platform for label-free biosensing. It has the advantage of being readily scalable to simultaneous readout of many PMCLs in array format, and allows integration …