研究目的
To develop a novel quencher-free fluorescent method for sensitive and specific detection of ochratoxin A (OTA) based on photoinduced electron transfer (PIET) between guanine and fluorophore, and to apply it in real samples like red wine.
研究成果
The developed quencher-free fluorescence method is highly sensitive and specific for OTA detection, with a low detection limit of 1.3 nM and successful application in red wine samples. It offers a simple, cost-effective alternative to traditional methods and has potential for broader applications in food safety and environmental monitoring.
研究不足
The method may have limitations in terms of specificity in highly complex matrices beyond red wine, and the need for optimization of aptamers for other toxins. Potential areas for optimization include further reducing detection limit and expanding to other sample types.
1：Experimental Design and Method Selection:
The method is based on the principle of photoinduced electron transfer (PIET) between a FAM-labeled OTA aptamer and a complementary strand (OTA-cAPT) containing guanines. In the absence of OTA, hybridization quenches fluorescence via PIET; in the presence of OTA, binding forms a G-quadruplex structure, maintaining fluorescence.
2：Sample Selection and Data Sources:
OTA, OTB, AFB1 toxins, and red wine samples were used. DNA probes were synthesized, and reagents were of analytical grade.
3：List of Experimental Equipment and Materials:
Includes F-2700 fluorescence spectrophotometer (Hitachi), DNA probes (Sangon Biotechnology), toxins (Pribolab, Yuanye), and various chemicals (Sinopharm Chemical Reagent).
4：Experimental Procedures and Operational Workflow:
Optimize conditions (e.g., aptamer concentration, reaction times), mix OTA with buffer and aptamer, incubate, add OTA-cAPT, measure fluorescence intensity. For selectivity, test with other toxins and metal ions. For real samples, filter and dilute red wine, spike with OTA, and measure.
5：Data Analysis Methods:
Fluorescence intensity measured at 520 nm, linear regression for calibration curve, detection limit calculated, and recovery rates determined.
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