Nucleic acid hybridization-based detection of pathogenic RNA using microscale thermophoresis

Matan Yosef Avivi, Noga Touitou, Hanan Rohana, Batia Lerrer, Yaron Shav-Tal, Avi Peretz, Haim Yosef Cohen

Research output: Contribution to journalArticlepeer-review

Abstract

Infectious diseases are one of the world's leading causes of morbidity. Their rapid spread emphasizes the need for accurate and fast diagnostic methods for large-scale screening. Here, we describe a robust method for the detection of pathogens based on microscale thermophoresis (MST). The method involves the hybridization of a fluorescently labeled DNA probe to a target RNA and the assessment of thermophoretic migration of the resulting complex in solution within a 2 to 30-time window. We found that the thermophoretic migration of the nucleic acid-based probes is primarily determined by the fluorescent molecule used, rather than the nucleic acid sequence of the probe. Furthermore, a panel of uniformly labeled probes that bind to the same target RNA yields a more responsive detection pattern than a single probe, and moreover, can be used for the detection of specific pathogen variants. In addition, intercalating agents (ICA) can be used to alter migration directionality to improve detection sensitivity and resolving power by several orders of magnitude. We show that this approach can rapidly diagnose viral SARS-CoV2, influenza H1N1, artificial pathogen targets, and bacterial infections. Furthermore, it can be used for anti-microbial resistance testing within 2 h, demonstrating its diagnostic potential for early pathogen detection.

Original languageEnglish
Article number105676
JournalJournal of Biological Chemistry
Volume300
Issue number3
DOIs
StatePublished - 1 Mar 2024
Externally publishedYes

Keywords

  • DNA
  • Doxorubicin
  • RNA
  • bacterial/viral infection
  • clinical diagnosis
  • microscale-thermophoresis
  • nucleic acids
  • pathogen detection
  • pathogen quantification

ASJC Scopus subject areas

  • Molecular Biology
  • Biochemistry
  • Cell Biology

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