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Aptamer-Based Target Detection Facilitated by a 3-Stage G-Quadruplex Isothermal Exponential Amplification Reaction
Published on: October 6, 2022
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A ratiometric fluorescent aptamer assay based on RCA amplification mult-G-quadruplex.
Xinyu Xie1, Weiling Li1, Zhiguang Suo2
1College of Food Science and Technology, Henan Key Laboratory of Cereal and Oil Food Safety and Nutrition, Henan University of Technology, Zhengzhou, 450001, China.
Analytical and Bioanalytical Chemistry
|August 27, 2025
Summary
A novel dual-signal assay detects Aflatoxin B1 (AFB1) by monitoring two fluorescence outputs. This ratiometric approach enhances accuracy by reducing false positives in AFB1 detection.
Area of Science:
- Biochemistry
- Analytical Chemistry
- Molecular Biology
Background:
- Single-signal detection assays are prone to false positives.
- Aflatoxin B1 (AFB1) is a significant food contaminant requiring accurate detection methods.
Purpose of the Study:
- To develop a ratiometric assay for Aflatoxin B1 (AFB1) detection.
- To leverage dual fluorescence signals for enhanced assay specificity and reduced false positives.
Main Methods:
- Designed a dual-signal assay utilizing aptamer-AFB1 binding and DNA strand interactions.
- Employed FAM (6-carboxyfluorescein) fluorescence quenched upon AFB1 binding to aptamer.
- Incorporated N-methylmesoporphyrin IX (NMM) fluorescence, dependent on G-quadruplex structure disruption.
Main Results:
- AFB1 binding to aptamer released a complementary strand, disrupting the G-quadruplex structure.
- Disruption of the G-quadruplex prevented NMM dye intercalation and fluorescence emission.
- A ratiometric signal based on FAM and NMM fluorescence intensity was established for AFB1 detection.
Conclusions:
- The developed dual fluorescence signal ratiometric assay offers a reliable method for AFB1 detection.
- This strategy provides a prospective approach for constructing dual-signal ratio assays for various analytes.
