Work

Mimicking a Cellular Crowding Environment for Enzyme-Free Paper-Based Nucleic Acid Tests at the Point of Care

Year: 2024

Type: article

Abstract: High Resolution Image Download MS PowerPoint Slide Point of care (PoC) nucleic acid amplification tests (NAATs) are a cornerstone of public health, providing the earliest and most accurate diagnostic method for many communicable diseases in the same location where the patient receives treatment. Communicable diseases, such as human immunodeficiency virus (HIV), disproportionately impact low-resource communities where NAATs are often unobtainable due to the resource-intensive enzymes that drive the tests. Enzyme-free nucleic acid detection methods, such as hybridization chain reaction (HCR), use DNA secondary structures for self-driven amplification schemes, producing large DNA nanostructures, capable of single-molecule detection in cellulo . These thermodynamically driven DNA-based tests have struggled to penetrate the PoC diagnostic field due to their inadequate limits of detection or complex workflows. Here, we present a proof-of-concept NAAT that combines HCR-based amplification of a target nucleic acid sequence with paper-based nucleic acid filtration and enrichment capable of detecting sub-pM levels of synthetic DNA. We reconstruct the favorable hybridization conditions of an in cellulo reaction in vitro by incubating HCR in an evaporating, microvolume environment containing poly(ethylene glycol) as a crowding agent. We demonstrate that the kinetics and thermodynamics of DNA–DNA and DNA–RNA hybridization is enhanced by the dynamic evaporating environment and inclusion of crowding agents, bringing HCR closer to meeting PoC NAAT needs.

Source: ACS Sensors

DOI: 10.1021/acssensors.4c00539

Authors: Jeffrey W. Beard, Samuel L. Hunt, Alexander Evans, Coleman Goenner, Benjamin L. Miller

Institutions: University of Rochester Medicine, University of Rochester

Language: English

Topic: Biosensors and Analytical Detection

Subfield: Biomedical Engineering

Field: Engineering

Domain: Physical Sciences

Open Access status: hybrid

Estimated APC paid: $4,500

Funders: University of Rochester, National Institute of Allergy and Infectious Diseases, National Institute of Biomedical Imaging and Bioengineering, National Science Foundation Graduate Research Fellowship Program