Assay development for pathogen detection at the point-of-need
Date
2020
Authors
Carrell, Cody S., author
Henry, Charles S., advisor
Farmer, Delphine K., committee member
McNally, Andrew, committee member
Reardon, Kenneth F., committee member
Journal Title
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Volume Title
Abstract
Infectious diseases are responsible for roughly one third of worldwide deaths, which disproportionately occur in low- and middle-income countries. Government health agencies recognize high quality diagnostics as a key tool to slow the spread and reduce the burden of disease in these countries. The same diagnostics that have minimized deaths from infectious disease in developed nations, however, cannot simply be implemented in all locations. Low- and middle-income countries lack the financial resources and infrastructure required to use the sophisticated instruments found in modern hospital laboratories. Instead of relying on current diagnostic technologies to reduce the burden of infectious disease, there is an urgent need to develop new technologies suited for the resource-limited settings they will be used in. The work described in this thesis aims to advance the capabilities of diagnostic sensors for use at the point-of-need. Microfluidic devices have been used for decades to perform complex analysis using compact devices with small sample and reagent volumes. Their portability and low-cost make them ideal candidates for analysis in resource limited settings, but their fabrication is tedious and expensive. To improve the fabrication process, Chapter Two of this thesis describes two methods for simplified 3D-printing of microfluidic devices. The 3D printer and resin used are inexpensive and commercially available and the fabrication process is not limited by the need to remove uncured resin from enclosed channels. Instead, open-faced channels in 3D-printed pieces were silanized and sealed to a secondary substrate. Common microfluidic devices including a droplet generator and herringbone mixer were created with the new fabrication method to demonstrate the strength of the seal and ability for the printer to create microfluidic channels. We envision this method being used for rapid prototyping and increased innovation in the field of microfluidic sensors. Traditional polymer microfluidics are limited in their usefulness in point-of-need situations because they require a pump to drive flow. Paper-based microfluidics use capillary action to drive flow instead of a pump and have emerged as an easy-to-use and inexpensive alternative to traditional microfluidics in situations where a power source is not available. However, paper-based microfluidics often suffer from poor analytical performance, and efforts to improve result in increased complexity. Chapter Three of this thesis describes a paper-based device that increases the sensitivity of a Salmonella assay while retaining ease-of-use. The device combines paper-pads for reagent storage with a 3D-printed rotational manifold to perform an enzyme-linked immunosorbent assay (ELISA). Typically, this assay requires dozens of complex pipetting steps, but the rotary device simplifies this process into four semi-automated steps. A detection limit of 440 colony forming units/mL was found using the paper-based device. As demonstrated in Chapter Three, common issues with paper-based microfluidics can be solved by integrating paper with other inexpensive components like 3D-printed polymer. In the final study in Chapter Four, we created a device to further simplify the steps of an ELISA using a combination of paper, polyester transparency film, and double-sided adhesive. The device, termed a disposable ELISA (dELISA), automatically performed the sequential reagent delivery and washing steps required for a traditional ELISA and require only two end user steps. The dELISA was then used to perform a serology assay for SARS-CoV-2 antibodies from whole-blood. The detection limit of the assay was 2.8 ng/mL for the dELISA, which was nearly identical to the detection limit found using a tradition well-plate assay (1.2 ng/mL).
Description
Rights Access
Subject
diagnostics
microfluidics
point-of-care
immunoassay
bioassay
paper-based