Studies On Highly Sensitive Methods For Key Factors Detection In Food Based On Nanoassembly And Immune Sensitizing

With the aid of nucleic acid moleculars, highly sensitive methods were developed for keyfactors (heavy metals ions, genetically modified DNA and melamine) detection in food,including Magnetic Resonance Imaging (MRI), Surface Enhanced Raman Spectroscopy(SERS), Colorimetric assay and Immuno-PCR. The main contents studied in this paper weredescribed as follows:1. A highly sensitive MRI sensor for the detection of Cu2+was established based on themagnetic nanoparticles assembly induced by the Cu2+-dependent ligation DNAzyme. TheDNA modified magnetic nanoparticles were proved by diameter change of particles beforeand after DNA conjugation, and the optimal concentration of DNAzyme was1μM. Atdifferent concentrations of Cu2+the magnetic nanoparticles assembly were characterized byTEM and DLS, which indicated that the aggregation level increased with the increase of Cu2+.For the quantitative analysis of Cu2+, a LOD of2.8nM was obtained in the Cu2+range of5nM to1000nM. In addition, this method has high specificity and excellent practicability.2. A DNA duplex composed of thymine-thymine (T-T) mispairs showed high selectivityand capturing capacity for Hg2+. Wih the property of this nucleic acid, an ultrasensitive SERSmethod for Hg2+detection was developed based on self-assembled gold nanoparticles chaininduced by T-Hg2+-T base pairs. In the presence of Hg2+, gold nanoparticles chains wereassembled successfully. A statistical analysis of the average number of AuNPs on chains ateach Hg2+concentration demonstrated that Hg2+concentration led to the chain elongation,resulting in SERS enhancement. The total E-field on the surface of chains measured usingcomputer simulations proved that the SERS enhancement arose from the high total E-feldintensity. In the Hg2+concentration range of0.001ng mL-1～0.5ng mL-1, a high sensitivitywith a LOD of0.45pg mL-1was obtained. Furthermore, this method with high specificity andpracticality provides enormous potential for trace amounts of Hg2+analysis.3. A colorimetric sensor based on the Ligation Chain Reaction (LCR) induced goldnanoparticles assembly was developed for genetically modified DNA detection. The more theconcentration of DNA, the greater the gold nanoparticles aggregates. This method wasrealized by the hybridization of probes and target DNA without the primer extension reaction.Using the UV-Vis signal as an indicator, a LOD of1.5aM was obtained. However, a LOD of0.1aM was obtained using DLS as an indicator. The specificity was good and the matrixinterference can be neglected.4. Combining the specifcity of competitive antigen–antibody reaction with theexponential amplifcation and quantitative effect of Real-Time quantitative PCR (RT-qPCR),an Immune-PCR for melamine (MEL) detection was developed. In the range of0.001pgg-1～10pg g-1, an excellent linear relationship between cycle threshold (Ct) and MELconcentration was established with a low LOD of0.3fg g-1. Compared with other methods,the sensitivity of this sensor showed a1000-fold improvement, and was below the strictestsafety limit of1ppm. Furthermore, the specifcity and the recovery in liquid milk samplesspiked with MEL were both excellent.