2021.7.27 K Byers Dissertation Final.pdf (7.07 MB)

Platforms and Molecular Mechanisms for Improving Signal Transduction and Signal Enhancement in Multi-step Point-Of-Care Diagnostics

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posted on 28.07.2021, 13:39 by Kaleb M. Byers

Swift recognition of disease-causing pathogens at the point-of-care enables life-saving treatment and infection control. However, current rapid diagnostic devices often fail to detect the low concentrations of pathogens present in the early stages of infection, causing delayed and even incorrect treatments. Rapid diagnostics that require multiple steps and/or elevated temperatures to perform have a number of barriers to use at the point-of-care and in the field, and despite efforts to simplify these platforms for ease of use, many still require diagnostic-specific training for the healthcare professionals who use them. Most nucleic acid amplification assays require hours to perform in a sterile laboratory setting that may be still more hours from a patient’s bedside or not at all feasible for transport in remote or low-resourced areas. The cold-chain storage of reagents, multistep sample preparation, and costly instrumentation required to analyze samples has prohibited many nucleic acid detection and antibody-based assays from reaching the point-of-care. There remains a critical need to bring rapid and accessible pathogen identification technologies that determine disease status and ensure effective treatment out of the laboratory.

Paper-based diagnostics have emerged as a portable platform for antigen and nucleic acid detection of pathogens but are often limited by their imperfect control of reagent incubation, multiple complex steps, and inconsistent false positive results. Here, I have developed mechanisms to economically improve thermal incubations, automate dried reagent flow for multistep assays, and specifically detect pathogenic antigens while improving final output sensitivity on paper-based devices. First, I characterize miniaturized inkjet printed joule-heaters (microheaters) that enable thermal control for pathogen lysis and nucleic acid amplification incubation on a low-cost paper-based device. Next, I explore 2-Dimensional Paper Networks as a means to automate multistep visual enhancement reactions with dried reagents to increase the sensitivity and readability of nucleic acid detection with paper-based devices. Lastly, I aim to create a novel Reverse-Transcription Recombinase Polymerase Reaction mechanism to amplify and detect a specific region of the Spike protein domain of SARS-CoV-2. This will allow the rapid detection of SARS-CoV-2 infections to aid in managing the current COVID-19 pandemic. In the future, these tools could be integrated into a rapid diagnostic test for SARS-CoV-2 and other pathogens, ultimately improving the accessibility and sensitivity of rapid diagnostics on multiple fronts.

Funding

National Science Foundation Small Business Innovation Research (SBIR) Program Grant 1720900

Purdue University Shaw Global Innovation Laboratory’s Innovations in International Development Award

Bill and Melinda Gates Grand Challenges Explorations program grant OPP1150806

Ross Fellowship from the Weldon School of Biomedical Engineering

History

Degree Type

Doctor of Philosophy

Department

Biomedical Engineering

Campus location

West Lafayette

Advisor/Supervisor/Committee Chair

Jacqueline C. Linnes

Additional Committee Member 2

Luis Solorio

Additional Committee Member 3

Ramses Martinez

Additional Committee Member 4

HyunDae Cho