Posner, Jonathan DMartin, Coleman2025-08-012025-08-012025Martin_washington_0250E_28027.pdfhttps://hdl.handle.net/1773/53450Thesis (Ph.D.)--University of Washington, 2025Nucleic acid diagnostics have advanced since PCR's first use in sickle cell and HIV diagnosis in the 1980s. PCR remains the gold standard for detecting SARS-CoV-2 and monitoring HIV viral load but is limited by resource-intensive requirements, making it impractical for low-resource or home settings. My research aims to adapt PCR’s strengths using isothermal nucleic acid amplification for rapid, low-cost diagnostics to support global health.I first present a novel assay combining RPA-based amplification with lateral flow detection, offering PCR-level sensitivity with LFA-like ease. It meets WHO SARS-CoV-2 detection standards, demonstrates high specificity, variant resilience, and uses a simple lysis method suitable for minimal devices. For HIV viral load monitoring, I developed a buffer-modified recombinase polymerase amplification assay on microfluidic chips using amplification nucleation site analysis (ANSA), where nucleation site counts correlate with nucleic acid input, enabling precise, mobile phone-compatible measurements. Finally, I describe a machine learning approach using a ResNet-18 model to analyze temporal ANSA data and predict DNA concentrations. Two models classify DNA by clinical groups or log-fold changes. This work supports robust, POC-suitable HIV diagnostics and establishes a platform for broader quantitative nucleic acid testing across global health settings.application/pdfen-USnoneComputer visionDiagnosticsHIVMicrofluidicsPoint of CareChemical engineeringBioengineeringChemical engineeringThesis