Sample Preparation for Point-of-Care Nucleic Acid Amplification Testing of Bloodborne Viruses

Abstract

Nucleic acid amplification tests (NAATs) provide high diagnostic accuracy for infectious diseases and quantitative results for monitoring viral infections. The majority of NAATs require complex equipment, cold chain dependent reagents, and skilled technicians to perform tests. This largely confines NAATs to centralized laboratories and can significantly delay appropriate patient care. Low-cost, point-of-care (POC) NAATs are especially needed in low- and middle-income countries (LMICs) for the detection of bloodborne infectious diseases, such as HIV, hepatitis B, hepatitis C, dengue, Zika, and Ebola viruses. The work presented in this dissertation describes novel sample preparation approaches for use in POC diagnostics for bloodborne pathogens. I developed paper-based isotachophoresis (ITP) systems for integrated on-chip detection and specialized nucleic acid purification of DNA and RNA. I also developed chemistries for inactivating endogenous blood ribonucleases and lysing viral envelopes, and I paired these chemistries with ITP for detecting HIV from human serum. My sample preparation methods use low-cost materials and reagents, and ITP uses an electric field to automate nucleic acid purification, reducing manual user steps. Ultimately, I believe this work may be used to develop POC NAATs for bloodborne pathogens in a low-cost format with minimal protocol complexity.