A universal biosensing platform for molecular diagnosis of emerging pathogens: From probe selection to clinical studies

Alafeef, Maha Mohammad Shehadeh

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https://hdl.handle.net/2142/120365 Copy

Description

Title

A universal biosensing platform for molecular diagnosis of emerging pathogens: From probe selection to clinical studies

Author(s)

Alafeef, Maha Mohammad Shehadeh

Issue Date

2023-04-19

Director of Research (if dissertation) or Advisor (if thesis)

Pan, Dipanjan

Doctoral Committee Chair(s)

Pan, Dipanjan

Committee Member(s)

• Bashir, Rashid
• Nie, Shuming
• Cunningham, Brian

Department of Study

Bioengineering

Discipline

Bioengineering

Degree Granting Institution

University of Illinois at Urbana-Champaign

Degree Name

Ph.D.

Degree Level

Dissertation

Keyword(s)

Biosensors, nanotechnology, diagnostic technologies, nanoparticles, infectious diseases

Abstract

COVID-19 continues to spread rapidly throughout the world, causing unprecedented disruption in modern society. While vaccines continue to be highly effective at preventing serious illnesses caused by this virus, new epidemiological data raised concerns about the arrival of new variants with potentially higher transmissibility. As these fast-emerging mutations fuel a fresh wave of infections, diagnostics continue to play a critical role in our efforts to contain and mitigate the pandemic. Rapid, easily accessible testing helps reduce the probability of mass outbreaks, as patients can be quickly identified and then self-quarantined. However, the current pandemic highlighted the limitations in existing analytical techniques that must be quickly adapted to stop the rapid spread of the virus. Diagnostics are applied to different patient populations in settings ranging from outpatient clinics and hospital intensive care units (ICUs) to point-of-care (POC) tests. Traditionally, the polymerase chain reaction (PCR) and the culture-based method served as the gold standard for the diagnosis of the pathogen, yet they are labor-intensive, slow, and not suitable for mass testing. The effective response to a pandemic of this magnitude would be the wide availability of assays that can identify the pathogen rapidly without compromising the test sensitivity. The accessibility of such tests will help in clarifying the etiology of the patient's illness, influence treatment modalities, and enable public health surveillance. We demonstrated that novel DNA probes can be designed and placed in various diagnostic platforms to satisfy the criteria outlined above for molecular detection of COVID-19. Diagnostic assays which we developed can be categorized into three main categories, i.e., i) laboratory-based techniques with comparable sensitivity to RT-qPCR but with much faster turnaround time, ii) point-of-care tests that offer the same speed and at the same time addresses the sensitivity issues with the widely available antigen tests, and iii) rapid tests that can be deployed at home for screening multiple viral infections at a time. Regardless of the sensor’s signal output, their high specificity is primarily achieved based on the use of single-stranded DNA or antisense oligonucleotides (ASO). In principle, every single sensor is constructed with this technology, has a single target, and will bind no other, even in a complex biological medium. While antibodies are quite versatile, they are prone to denaturation and cannot recognize every analyte. We engineered nucleic acid probes that interact with the genetic sequences of the virus irrespective of its ongoing mutations. The probes target a specific segment of the nucleocapsid phosphoprotein (N) gene of SARS-CoV-2, which does not mutate among the known variants, with high binding efficiency. To achieve sensitive detection of pathogens, two distinct characteristics of nanomaterials, i.e., optical (plasmonic and scattering) and electrochemical were employed. The signal output relies upon the changes that result from the direct interaction of SARS-CoV-2 RNA and a derivatized nucleic acid probe. The developed platform supports the interchangeability of the synthetic oligonucleotide responsible for viral detection allowing for rapid adaptation to new pathogens. Indeed, the diagnostic platform should be able to rival the deployment of PCR and other nucleic acid amplification tests (NAAT) in the event a new genomic sequence of a new pathogen is available. To date, in addition to COVID-19, we have demonstrated that the approach can be used for multiplexing other pathogens, e.g., Flu A virus.

Graduation Semester

2023-05

Type of Resource

Thesis

Copyright and License Information

Copyright 2023 Maha Alafeef

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