Aptamer-sensing Analysis Based On Enzymatic Cascade Amplification And DNA-Scaffold

With the continuous development and improvement of the analytical science, various analysis and detecting process are increasingly needed the help of biosensing technology to achieve the required information. At present, the biosensing technology has been developed rapidly due to many merits such as high sensitivity, good selectivity, fast response and low cost. However, greater biosensing techniques are required with the development of scientific research. Therefore, accuracy strategies with high sensitivity and selectivity are attracte more concerns in chemistry analytical field. Enzymatic cascade amplification and DNA-scaffold technique were proposed to detect L-histidine and mental ions. The details are mainly performed as following:A novel DNAzyme based enzymatic cascade strategy has been devised for the exponential amplified detection of L-histidine. This is the first attempt to carry out the combination of the DNAzyme and the polymerase/endonuclease reaction cycles leading to the direct activation of an enzymatic cascade. In the presence of L-histidine, the enzyme strand carries out catalytic reactions to hydrolytic cleavage of the substrate strand at the scissile rA. The cleaved 11-mer primer with the ribonucleotide rA hybridizes with template strand and triggers the Klenow Fragment polymerization. Nb.BbvCI endonuclease cuts the double-stranded niking site and thus opens a new site for a new replication. The enzymatic cascade regenerates the completely dsDNA to initiate another cycle of nicking, polymerization and displacement. Finally the fluorescence dye, SQ inserts into the DNA double helix to generate a distinguishable fluorescence enhancement. A good nonlinear correlation (R=0.9994) between fluorescence intensity and the logarithm of the L-histidine concentration over the range 50 nM～1.0 mM. The detection limit as low as 6.34 nM. This DNAzyme based enzymatic cascade strategy shows great potential for future use in clinical tests owing to its highthroughput, isothermal, and cost-efficient features.We herein report a simple, sensitive and rapidly fluorescent sensor based on DNA stabilized silver nanoclusters (DNA-AgNCs) for the determination of mercury ions(Hg2+). Ascribed to the strong affinity of silver ion (Ag+) with cytosine-cytosine base pairs, the stable C-Ag+-C complexes are formed. After reduced by sodium borohydride (NaBH4), the synthesised DNA-AgNCs exhibited strong fluorescence. While upon the addition of Hg2+, DNA-AgNCs shows a dramatic decrease in fluorescence which devoted to the metallophilic interaction between Hg2+ with Ag+ that present in NCs. The present assay allows for the sensing of Hg2+ in the range of 5.0 nM to 8.0μM with detection limit as low as 1.57 nM. This proposed method may expand the application of DNA-AgNCs in the field of heavy mental monitoring.C-rich hairpin probes involving graphite oxide (GO) was developed for detection of silver ions. Silver ions are specific recognized by C-rich hairpin probes to form stable C-Ag+-C complexes. Thus the C-rich hairpin probes were transfered to double-strand DNA (dsDNA). Intercalated dye SYBR Green1 (SG) was employed as reporter for the readout of the conversation of the C-Ag+-C complexes. The GO reduces the background as a quencher to the minmum to obtain the higher signal background ratio. Under the optimized experimental conditions, the fluorescence intensity showed a good linear reponse to the logarithm of the Ag+ concentration in the range of 20 nM～2.0μM. The detection limit for Ag+ was 6.47 nM. This proposed method shows and also excellent selectivity, with little interference from other common metal ions. The recovery was found to vary from 93.37% to 101.2% indicating the application potential of this assay for real samples.