Nucleic acid hybridization-based detection of pathogenic RNA using microscale thermophoresis.
J Biol Chem
; 300(3): 105676, 2024 Mar.
Article
en En
| MEDLINE
| ID: mdl-38278326
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.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
ARN
/
Técnicas Microbiológicas
/
Técnicas de Diagnóstico Molecular
/
Ensayos Analíticos de Alto Rendimiento
/
Hibridación de Ácido Nucleico
Tipo de estudio:
Diagnostic_studies
Límite:
Humans
Idioma:
En
Revista:
J Biol Chem
Año:
2024
Tipo del documento:
Article
País de afiliación:
Israel