Ultrasensitive Detection Of Ochratoxin Research Articles

Recognition-Activated Primer-Mediated Exponential Rolling Circle Amplification for Signal Probe Production and Ultrasensitive Visual Detection of Ochratoxin A with Nucleic Acid Lateral Flow Strips.

We proposed a visual strategy for rapid and ultrasensitive detection of ochratoxin A (OTA) by integration of primer-mediated exponential rolling circle amplification (P-ERCA) with a designed nucleic acid lateral flow strip (LFS). The recognition component was preimmobilized in the tube by hybridization between the immobilized functionalized aptamer and complementary ssDNA. Recognition of OTA induces the release of complementary ssDNA from the tube, which will also act as the primer of the designed P-ERCA. Three nicking sites on the template P-ERCA could contribute to the production of enormous signal probes based on the simultaneous amplification-nicking model, which can be visually measured directly with the constructed nucleic acid LFS. Importantly, the nicked signal probe can also act as the trigger of the new-round RCA, achieving exponential growth of signal probes for measurement and signal enhancement. Taking advantage of the extraordinary amplification efficiency of P-ERCA and the simplicity of LFS, this P-ERCA-LFS method demonstrates ultrasensitive detection of OTA with a visual limit of detection as low as 100 fg/mL for qualitative screening and a limit of detection of 35 fg/mL for semiquantitative analysis. This designed strategy could also be utilized as a universal method for detection of other chemical analytes with the replacement of the aptamer for recognition, and the nucleic acid LFS unit could also be a useful protocol for direct ssDNA analysis.

A novel magneto-gold nanohybrid-enhanced lateral flow immunoassay for ultrasensitive and rapid detection of ochratoxin A in grape juice

Conventional gold nanoparticle-based lateral flow immunoassay (LFIA) usually suffers a huge challenge in measuring target concentration in food matrices with dark color because of its poor resistance to the background matrix and color interference. To address this issue, we first report a novel bifunctional magneto-gold nanohybrid (MGNH) for the simultaneous magnetic separation and colorimetric target sensing by integrating MGNHs into LFIA. Under optimum conditions, an ultrasensitive detection of ochratoxin A (OTA) in grape juice was achieved with a limit of detection at 0.094 ng mL−1. The average recoveries of this MGNH-LFIA ranged from 92.31% to 108.97% with a coefficient of variation of below 12%. The excellent selectivity of our MGNH–LFIA against OTA was demonstrated. Besides, our MGNH–LFIA is comparable to liquid chromatography coupled with mass spectrometry in terms of accuracy, reproducibility, and practicability. The designed MGNH-LFIA platform is readily extended for improving other small molecule detection in food samples.

Ultrasensitive detection of ochratoxin A based on biomimetic nanochannel and catalytic hairpin assembly signal amplification

In this paper, an ultrasensitive nanochannel sensor has been proposed for label-free Ochratoxin A (OTA) assay in combination with graphene oxide (GO) and catalyzed hairpin assembly (CHA). The high-performance sensor is segmented into two parts. One is composed of graphene oxide (GO) and DNA probes. In the presence of target OTA, OTA works as a catalyst to trigger the self-assembly pathway of the two probes and initiate the cycling of CHA circuits, which results in numerous double-stranded DNAs (dsDNA) in solution. The excess ssDNA probes are removed by GO. The other part is composed of biomimetic nanochannel coated with polyethyleneimine (PEI) and Zr4+, which can quantify the concentration of OTA by detecting the dsDNA in solution. The nanofluidic device has a detection limit of as low as 6.2 pM with an excellent selectivity. The nanochannel based assay was used to analyse food samples (red wine) with satisfied results. Thus, the proposed analytical method will provide a new approach the detection of OTA and can be applied for quality control to ensure food safety.

Rolling Chain Amplification Based Signal-Enhanced Electrochemical Aptasensor for Ultrasensitive Detection of Ochratoxin A

A novel electrochemical aptasensor is described for rapid and ultrasensitive detection of ochratoxin A (OTA) based on signal enhancement with rolling circle amplification (RCA). The primer for RCA was designed to compose of a two-part sequence, one part of the aptamer sequence directed against OTA while the other part was complementary to the capture probe on the electrode surface. In the presence of target OTA, the primer, originally hybridized with the RCA padlock, is replaced to combine with OTA. This induces the inhibition of RCA and decreases the OTA sensing signal obtained with the electrochemical aptasensor. Under the optimized conditions, ultrasensitive detection of OTA was achieved with a limit of detection (LOD) of 0.065 ppt (pg/mL), which is much lower than previously reported. The electrochemical aptasensor was also successfully applied to the determination of OTA in wine samples. This ultrasensitive electrochemical aptasensor is of great practical importance in food safety and could be widely extended to the detection of other toxins by replacing the sequence of the recognition aptamer.