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Updated: Aug 6, 2026
Probe-based Real-time PCR Approaches for Quantitative Measurement of microRNAs
Published on: April 14, 2015
Self-assembled multifunctional hairpin probe for ultrasensitive and mismatch-selective miRNA detection
Baoqiang Chen1, Yiping Fan2, Qi Wang3
1Provincial Key Laboratory of Multimodal Perceiving and Intelligent Systems, Engineering Research Center of Intelligent Human Health Situation Awareness of Zhejiang Province, Jiaxing Key Laboratory of Molecular Recognition and Sensing, College of Biological and Chemical Engineering, Jiaxing University, Jiaxing, Zhejiang, 314001, China.
A new molecular probe, assembled multifunctional hairpin probe (A-MF-HP), offers sensitive and stable microRNA detection. This tool enables robust diagnostics in biological samples, even in human serum.
Area of Science:
- Molecular Diagnostics
- Biochemistry
- Nucleic Acid Chemistry
Background:
- Detecting microRNAs (miRNAs) in biological samples is challenging due to sensitivity, specificity, and stability issues.
- Existing methods often require multiple components and are susceptible to degradation in complex environments like serum.
Purpose of the Study:
- To develop a novel, single-molecule probe for sensitive, specific, and stable detection of miRNAs.
- To create a tool that integrates target recognition, signal amplification, and fluorescence reporting autonomously.
Main Methods:
- Introduction of a bidirectional palindromic assembled multifunctional hairpin probe (A-MF-HP).
- The probe utilizes autonomous self-assembly into a nuclease-resistant structure.
- miRNA-21 recognition triggers a cascade signal amplification via polymerase extension, nicking, and strand displacement.
Main Results:
- The A-MF-HP platform achieved femtomolar sensitivity (10^-15 M) with a six-order dynamic range.
- Demonstrated single-nucleotide mismatch discrimination and negligible cross-reactivity.
- The probe maintained functionality in 10% human serum for 12 hours and showed elevated fluorescence in lung cancer patient samples.
Conclusions:
- A-MF-HP offers a robust, single-probe solution for miRNA detection, overcoming limitations of multi-component assays.
- The technology provides biostability, multifunctionality, and autonomous amplification for miRNA profiling.
- This novel strategy presents a powerful diagnostic tool for biomedical and clinical applications.
