Nanographite-based fluorescent biosensor for detecting microRNA using duplex-specific nuclease-assisted recycling

Area of Science:

• Biochemistry
• Nanotechnology
• Molecular Biology

Background:

• MicroRNA (miRNA) detection is crucial for disease diagnosis and understanding biological processes.
• Existing miRNA detection methods often face challenges with sensitivity and specificity.
• Development of novel, highly sensitive biosensors is essential for advancing miRNA research.

Purpose of the Study:

• To develop a novel nanographite (NG)-based fluorescent biosensor for sensitive and specific microRNA (miRNA) detection.
• To utilize duplex-specific nuclease (DSN) for signal amplification in the biosensing system.
• To evaluate the biosensor's performance, including detection limit, linear range, and specificity.

Main Methods:

• A nanographite (NG) material was employed as the platform for a fluorescent biosensor.
• A double carboxyfluorescein (FAM)-labelled probe (DFP) was designed to be complementary to the target miRNA.
• Duplex-specific nuclease (DSN)-assisted signal amplification and p-stacking interactions were utilized for fluorescence modulation.

Main Results:

• The biosensor exhibited fluorescence quenching in the absence of target miRNA due to NG adsorption of the DFP probe.
• Fluorescence was restored in the presence of target miRNA, with signal amplification attributed to DSN-mediated target recycling.
• The biosensor achieved a detection limit of 10 pmol/L for miRNA, with a linear correlation observed between 50 pmol/L and 5 nmol/L.
• The method demonstrated specificity, distinguishing let-7b from other let-7 miRNA family members, and was validated in a sample assay.

Conclusions:

• The developed nanographite-based fluorescent biosensor is a novel and highly sensitive tool for miRNA detection.
• The DSN-assisted signal amplification strategy significantly enhances detection sensitivity.
• This biosensor holds great potential for applications in biochemical research, disease diagnosis, and therapeutic monitoring.