"DSN-mismatched CRISPRâ³sensor for highly selective and sensitive detection of under-expressed miR-let-7a.
Anal Chim Acta
; 1295: 342273, 2024 Mar 22.
Article
en En
| MEDLINE
| ID: mdl-38355234
ABSTRACT
Several microRNAs (miRNAs) are expressed at lower levels in specific tumors, e.g., miR-let-7a in non-small cell lung cancer (NSCLC). This makes it challenging to analyze their lower abundance versus specifically elevated miRNAs. Here, we describe a novel fluorescent biosensor for the highly selective and sensitive detection of miR-let-7a constructed by combining miRNA screening assisted by a duplex-specific nuclease (DSN) with CRISPR-Cas12a system signal amplification. We meticulously designed a mismatch in the first three to four bases at the 5'-end of the capture DNA to improve the signal-to-noise ratio of the CRISPR-Cas12a system. Within this "DSN-mismatched CRISPR" fluorescence strategy, miR-let-7a was accurately screened by DSN-assisted cleavage, and the mismatched capture DNA unbound to target miRNA could trigger the CRISPR-Cas12a system to produce a mass of trans-cleave fluorescence signals. This "turn-off" approach was suitable for detecting decreased levels of miRNAs. This approach can not only discriminate the single-base mismatched let-7 family but also reach a limit of detection at 64.17 fM as well as be quantified from 100 fM to 500 pM. The miR-let-7a levels were then measured in clinical serum samples from healthy volunteers and patients with NSCLC. This study holds promise for the development of a universal under-expressed miRNA assay for early diagnosis and treatment of cancers.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Técnicas Biosensibles
/
Carcinoma de Pulmón de Células no Pequeñas
/
MicroARNs
/
Neoplasias Pulmonares
Tipo de estudio:
Diagnostic_studies
/
Screening_studies
Límite:
Humans
Idioma:
En
Revista:
Anal Chim Acta
Año:
2024
Tipo del documento:
Article
Pais de publicación:
Países Bajos