A Novel and Label-Free Chemiluminescence Detection of Zearalenone Based on a Truncated Aptamer Conjugated with a G-Quadruplex DNAzyme

Abstract

Zearalenone (ZEN), one of the most frequently occurring mycotoxin contaminants in foods and feeds, poses considerable threat to human and animal health, owing to its acute and chronic toxicities. Thus, rapid and accurate detection of ZEN has attracted broad research interest. In this work, a novel and label-free chemiluminescence aptasensor based on a ZEN aptamer and a G-quadruplex DNAzyme was constructed. It was established on a competitive assay between ZEN and an auxiliary DNA for the aptamer, leading to activation of the G-quadruplex/hemin DNAzyme and subsequent signal amplification by chemiluminescence generation after substrate addition. To maximize the detection sensitivity, numerous key parameters including truncated aptamers were optimized with molecular docking analysis. Upon optimization, our aptasensor exhibited a perfect linear relationship (R2 = 0.9996) for ZEN detection in a concentration range of 1–100 ng/mL (3.14–314.10 nM) within 40 min, achieving a detection limit of 2.85 ng/mL (8.95 nM), which was a 6.7-fold improvement over that before optimization. Most importantly, the aptasensor obtained a satisfactory recovery rate of 92.84–137.27% and 84.90–124.24% for ZEN-spiked wheat and maize samples, respectively. Overall, our label-free chemiluminescence aptasensor displayed simplicity, sensitivity, specificity and practicality in real samples, indicating high application prospects in the food supply chain for rapid detection of ZEN.

Truncations and in silico Docking to Enhance the Analytical Response of Aptamer-Based Biosensors

“…DeepFoldRNA is a web-based software that models tertiary structures using deep learning techniques, which has shown improved performances on benchmark datasets like Rfam families or RNA-Puzzle experiments with root mean square deviation (RMSD) averages of 2.69 Å in comparison to other models. To use DeepFoldRNA, as others have prior to us, [59][60][61][62][63] we transposed the glucose aptamer sequence into RNA format. From the 10 predictions generated by the algorithm, we picked the one with the lowest RMSD and then swapped the uracils for thymines in the structure using the web-based tool, x3DNA, 54 prior to removing the 2'-OH on the ribose backbone using PyMol.…”

Smartphone-Based Chemiluminescence Glucose Biosensor Employing a Peroxidase-Mimicking, Guanosine-Based Self-Assembled Hydrogel

“…Conversely, the guanosine/hemin CL hydrogel provided glow-type, longlasting photon-emission kinetics and the maximum CL was observed several minutes after To further investigate the CL response of the hydrogel to H 2 O 2 , we compared its CL kinetic profile to that of a ssDNA designed hemin/G-quadruplex DNAzyme. Indeed, hemin/DNA-based G-quadruplex structures prepared employing guanine-rich ssDNA sequences are widely used as peroxidase-like DNAzymes in CL biosensors [57]. As shown in Figure 3, upon addition of a CL cocktail containing 10 mmol L −1 luminol and 1.0 mmol L −1 H 2 O 2 in carbonate buffer, the ssDNA designed DNAzyme displayed a fast photon emission kinetics, in which the CL emission reached its maximum intensity in few seconds, then decayed to background signal in about ten minutes.…”

Towards a lab-on-chip sensor: chemiluminescence-based DNA-nanomachine for nucleic acid detection at zeptomolar concentrations

“…Previous studies have already explored CL methods based on peroxidase-like DNAzymes using split G-4 probes for nucleic acid detection. These studies showcased sensitivity ranging from nanomolar to femtomolar concentrations [10][11][12][13][14] and successful discrimination of single nucleotide substitution (SNS) [10,12]. The sensitivity of the presented systems may be affected by the number of immobilized sensory oligonucleotides on the flowcell surface or by inclusion of additional external manipulations before introducing the sample to the CL detection system.…”