<p>As of 2021, 38.4 million people worldwide are living with Human Immunodeficiency virus (HIV), with eastern and southern Africa having the highest prevalence. The efficacy of treatment is determined by identifying acute HIV infections (AHI) and prompting early antiretroviral therapy (ART) initiation to achieve viral suppression and reduce the risk of transmission. Existing rapid tests that detect host antibodies are affected by long seroconversions which allow the viruses to remain undetected until long after infection. On the contrary, highly sensitive nucleic acid amplification test (NAAT) based assays, serving as the gold standard for detection are restricted by their long turnaround time and high cost of implementation thus, restricting their use in low resource settings. Further, drug resistance cases and patient non-compliance to treatment may lead to HIV progression to aids; therefore, effective viral load monitoring is a critical component in the HIV care continuum. To address the gaps in viral load monitoring and early HIV detection, I propose to develop assays for handheld self-test platforms to detect low concentrations of HIV via two different approaches: 1) I will optimize an existing NAAT - based assay to semi-quantitatively detect HIV particles that were spiked in clinical samples and 2) I will Investigate the binding kinetics between HIV p24 antigen and Anti-HIV-1 p24 Antibody using the principle of Bio-layer Interferometry. Thus, I will lay the foundation for the development of a novel and highly sensitive p24 detection assay. Overall, this work will enable detection of ahi detection as well as support people living with HIV (PLHIV) management, all while remaining connected to healthcare and provider support. </p> <p><br></p>

<p>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.</p> <p>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.</p>