An Aptamer-based Fluorescence Assay for Ochratoxin A

A novel analytical method for the detection of Ochratoxin A (OTA) is established based on the aptamer recognition and fluorescent probe technology. The present method is developed based on the fact that when the immobilized aptamer bonds to the target OTA, it can induce the conformation change of aptamer and result in the dissociation of the FAM-tagged complementary DNA chain from aptamer, finally leading to the fluorescent signal change. Based on it, OTA can be quantified. All the condition factors affecting the performance of the present method are investigated. The results show if the avidin concentration coated on the microplate is 25 mg L−1, the aptamer concentration is 50 nM, the FAM-tagged complementary DNA chain concentration is 150 nM, the binding buffer solution is chosen as 10 mM HEPES, pH 7.0 (contains 120 mM NaCl, 5 mM KCl, 20 mM MgCl2, and 20 mM CaCl2), and the binding reaction is conducted at 45°C for 40 min, the optimal analytical performance can be achieved for the present work. Under the optimal conditions, the linear range for the OTA concentration detection is 2.0 × 10−8-1.0 × 10−5 g L−1 with a detection limit of 1 × 10−8 g L−1. The RSD is 2.6% for 11 parallel measurements of 1 × 10−6 g L−1 OTA. Meanwhile, the present method is highly selective for OTA and easy to be operated. It has been successfully applied to measure OTA content in real samples.