Construction of two fluorescence aptasensors with RecJf exonuclease-induced signal amplification for detecting aflatoxin B1.

Abstract

Aflatoxin contamination, especially aflatoxin B1 (AFB1), is a major global health concern. There is an urgent need to develop rapid, highly sensitive and specific sensing platforms. In this study, two fluorescence aptasensors were developed for AFB1 detection. The first used quantum dots (QDs) combined with aptamers as molecular recognition elements, along with RecJ_f exonuclease amplification and Fe₃O₄ nanoparticles for magnetic separation. In the absence of AFB1, the aptamer stably hybridizes with the cDNA-QDs conjugate, thereby shielding its 5' end from cleavage by RecJ_f exonuclease. Upon AFB1 binding, the duplex dissociates, exposing the aptamer's 5'-single-stranded region to RecJ_f-mediated hydrolysis-resulting in AFB1 release and enabling catalytic recycling and cascade signal amplification. It achieved a detection range of 0.01-100 ng mL^-1 and a limit of 2.2 pg mL^-1 (3σ/s). To further reduce interference and improve sensitivity, a second sensing system based on upconversion nanomaterials (UCNPs) was developed. This system, also using RecJ_f exonuclease amplification, achieved a broader range of 0.001-100 ng mL^-1 and a lower detection limit of 0.56 pg mL^-1. The QDs-based system required 90 min for detection, whereas the UCNPs-based system achieved detection in just 10 min. The two detection methods, particularly the UCNPs based one, enable specific identification of AFB1. Both aptasensors were successfully applied to the determination of AFB1 in spiked wheat flour and corn flour samples, with recoveries ranging from 89% to 111.6%, demonstrating their applicability to AFB1 analysis in cereal products. By replacing the aptamer, the proposed aptasensor can be further extended for the detection of other biological or chemical targets.