Molecular aptamer beacon-based dual-mode exponential isothermal amplification for microRNA detection

Area of Science:

• Biotechnology
• Molecular Biology
• Analytical Chemistry

Background:

• Current microRNA (miRNA) detection methods face limitations like low abundance and short sequences, hindering rapid clinical use.
• The gold standard for miRNA detection is not suitable for low-cost, rapid clinical diagnostics.

Purpose of the Study:

• To develop a novel, rapid, and cost-effective method for microRNA (miRNA) detection in biological samples.
• To establish a one-pot assay for analyzing and detecting miRNAs with improved sensitivity and efficiency.

Main Methods:

• Developed a one-pot method: molecular aptamer beacon-utilized exponential isothermal amplification (MABEXIA).
• Utilized both fluorescence and chemiluminescence detection platforms for miRNA analysis.
• Employed DNA polymerase (Klenow Fragment) and nicking endonuclease (Nb.BbvCI) for signal amplification in the fluorescence method.
• Incorporated streptavidin (SA) aptamer and SA-coated magnetic beads for chemiluminescence detection.

Main Results:

• The MABEXIA method demonstrated good detection performance in 1% human serum.
• The fluorescence platform generated significant double-stranded (ds) DNA products upon target miRNA hybridization.
• The chemiluminescence approach exhibited a good linear relationship (50 fM to 50 pM) and detection ability comparable to the gold standard.

Conclusions:

• MABEXIA offers a promising alternative for sensitive and efficient miRNA detection.
• The developed method, particularly the chemiluminescence approach, meets the requirements for rapid and low-cost clinical miRNA analysis.
• This technique has the potential to overcome the limitations of existing miRNA detection methods.