Enzyme-assisted nanopore sensor for sensitive and multiplexed detection of microRNAs

Area of Science:

• Biotechnology
• Nanotechnology
• Molecular Biology

Background:

• MicroRNAs (miRNAs) are critical in disease but difficult to detect due to low abundance.
• Multiplexed detection of miRNAs requires high sensitivity and specificity.

Purpose of the Study:

• To develop an enzyme-assisted nanopore sensor for simultaneous and ultrasensitive detection of multiple miRNAs.
• To improve sensitivity and specificity for low-abundance miRNA detection.

Main Methods:

• Utilized streptavidin-coated magnetic beads with capture DNA (cDNA) for miRNA hybridization.
• Employed alkaline phosphatase (ALP)-labeled detection DNA and ALP-catalyzed dephosphorylation of peptide probes.
• Integrated nanopore sensing with cucurbit [7]uril for signal generation and magnetic separation for background reduction.

Main Results:

• Achieved simultaneous detection of three target miRNAs with femtomolar-level limits of detection.
• Demonstrated a broad dynamic range from 10.0 fM to 10.0 nM.
• Showcased significantly improved sensitivity for low-abundance miRNAs through enzymatic amplification and signal enhancement.

Conclusions:

• The enzyme-assisted nanopore sensor provides a robust platform for precise miRNA analysis.
• This technology holds significant potential for early disease diagnosis by enabling sensitive detection of miRNAs.