Electrokinetic platforms for the enrichment and detection of biological targets
Date
2023-12
Authors
Osman, Sommer
Major Professor
Advisor
Anand, Robbyn K
Lee, Young-Jin
Claussen, Jonathan
Pandey, Santosh
Anderson, Jared
Committee Member
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Altmetrics
Abstract
The COVID-19 pandemic was exacerbated by a lack of available testing, with most samples being sent to centralized labs that suffered from long wait times and a lack of available testing materials. There is a clear need for better testing at the point-of-care (POC), the place in which patients must receive care, such as at the clinic, home, or even at screening sites, including airports and schools. If widespread testing had been implemented during this critical time, public health measures, such as self-quarantine and contract tracing, would have been more effective in slowing transmission of the virus. Above all, the use of POC testing has been proven to improve clinical outcomes for patients.
To meet this gap, many researchers have focused on the development of antigen-based testing instead of gold-standard nucleic acid-based testing (e.g., RT-PCR). Antigen-based testing meets much of the ASSURED criteria, a framework developed by the World Health Organization (WHO) that outlines the ideal POC test; they are more affordable, typically use fewer reagents, and, most importantly, achieve results rapidly, often within an hour. These POC immunoassays can be realized in several platforms, including paper-based analytical devices (PADs) and lab-on-a-chip (LOC). PADs were one of the first POC diagnostics to emerge and are unique due to the special properties of paper. PADs have numerous advantages, including the ability to easily store necessary reagents and pretreatments, the elimination of external pumps for sample flow, simple fabrication, and portability. However, PADs are plagued with poor sensitivity in comparison to standard laboratory techniques. In contrast, LOC technologies have historically been much more sensitive and reproducible, comprised of PDMS/glass with operation on the micro- or nanoscale. A major challenge of LOC devices is that they often rely on bulky equipment to operate, and are expensive and complicated to fabricate, which hinders mass production. Moreover, very few examples of LOC devices currently used for clinical-based testing exist.
Electrokinetics presents solutions to the challenges associated with the sensitivity and deliverability of POC testing. It is the movement of charged particles and fluids under an electric field and has been successfully leveraged for both PADs and LOC technologies. The most recently developed technique, ion concentration polarization (ICP), is of special interest because of its attained high enrichment factors and sustained preconcentration of analytes over long periods of time. ICP has been demonstrated in biofluids and often requires no sample pretreatment or specialized buffer systems, resulting in the selective transport of charged species. The underlying mechanism of ICP—surface conductance dominance over bulk conductance—has been used for ultra-sensitive sensing of biological targets and, notably, achieved using commercially available materials.
In this dissertation, several electrokinetic strategies have been designed to improve immunoassay sensitivity, in the context of deliverability, with the goal of creating diagnostics that are more amenable to the POC. The first chapter addresses the instability of focused protein plugs by integrating a packed bed of microbeads in an ICP-PAD. Evaluation of the microbead diameter's impact on parameters, such as average and maximum enrichment factor, as well as the retained fraction of the plug, reveals the optimum configuration for stable preconcentration of protein analytes. The second chapter details a LOC device for the enrichment and label-free, non-optical detection of a bead-based immunoassay by microscale surface ion conduction (µSIC). Several features of the sensor are explored, including a direct lateral flow immunoassay, an optional preconcentration step, and the ability for multiple interrogations to obtain kinetic information. The third chapter documents the µSIC's sensor adaption to the detection of whole virus particles, specifically SARS-CoV-2 virions. The sensor is successfully challenged in a biofluid for the first time, detailing a strategy that would overcome the heterogeneity of saliva and patient-to-patient variability for testing. The advantages of these platforms for POC testing include versatility with demonstrated plug-and-play capabilities, simple fabrication, improved sensitivity, rapid results, and microliter sample sizes. We envision the works described herein taking the next steps towards POC implementation to equip clinicians and patients more quickly with the information they need to make better-informed healthcare decisions.
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dissertation