Detection Of PDGF-BB And MRNA Based On The Synthesis Of Silver Nanoclusters

The fluorensence materials has been gradually wide applied in the area of bio-analysis.But classic fluorensence materials often suffered from various shortages while applied in actual application.Typical fluorensence materials mainly include the organic dyes and QDs.The high photostability of organic dyes is poor and its fluorensence is easy to be influenced by environment factors.The QDs possess large size and biological toxicity,which limited its application in the bio-cell imaging and biolabels.The silver nanocluster,as a novel fluorensence material,possess advantages such as good light stability,high quantum yield,small size,low biological toxicity and adjustable fluorensence transmission wavelength,which leads to a good application vista in various areas such as bioimaging,biosensing and biolabeling.In this thesis,the silver nanocluster was applied as Fluorensence labels.With the assistance of analytical techniques,such as Catalyzed Hairpin Assemby?CHA?,Proximity Ligation Assay?PLA?and Rolling circle amplification?RCA?etc,the detection for PDGF-BB and mRNA was realized.This paper mainly includes three aspects as following:?1?A simple and high-sensitive method for PDGF-BB detection is designed by applying the DNA-AgNCs as fluorensence labels.The existence of target protein PDGF-BB will caused the Configuration transformation of designed aptamer probe and generated stable aptamer-protein composite and further induce the Polymerization and Enzymatic Reaction to generate large amounts of gold nanocluster template.Furthermore,based on the chemical reduction reaction of AgNO₃ and NaBH₄ and adpoting the generated sliver nano clusters as template,the fluorensence-active DNA-AgNCs could be synthesized.The high-specificity interaction between aptamer and PDGF-BB provided high specifity for this detection method.Besides,the fluorensence of the DNA-AgNCs could be enhanced by G-rich DNA chains,which could further enhance the detection sensivity?0.06 nM?.Besides,this design is only based on the nuclear acid hybridization and Enzyme amplification amplification technology,thus,it could be applied for detecting other bio molecules by changing the aptamer sequences.?2?Up to now,the oligonucleotide-encapsulated sliver nanoclusters have been widely applied in bio-analysis.In this work,we developed a new method to encapsulate synthesize the DNA-AgNCs adopting cytosine-rich long chain DNA.Based on the TdT-polymerized long chain C-rich DNA and the Exo III assistanced signal amplification strategy,a label-free fluorensence sensing platform is constructed.The PDGF-BB was applied as the target.In this strategy,the specifically designed aptamer S1 could combine with the PDGF-BB,which caused by the proximity of the aplamer and intoduce the Proximity Ligation Assay to hybridize with the complementary probe S2.The Exo III will resecte the probe S2.As a result,the Polymerization cycle amplification end will release short DNA Chains and induce next polymerization cycle.Under the existence of TdT and dCTP,the long chain C-rich DNA could be synthesized as the growth template of fluorensence sliver nanoclusters to realize label-free PDGF-BB detection.This method possess high sensivity and low cost with a detection limit as low as 0.3 pM.Based on this nover strategy,our work proved that the TdT-based DNA-AgNCs synthesis strategy can be applied as multi-function and effective bio-sensing platform for bio molecules detection.?3?In order to avoid the complex interactions between the QDs and G-quadplex/hemin,the DNA-AgNCs,whic posses advantage such as easy for synthesis light stability,low Biological toxicity and especially the consistency between the Donors and receptors template attracted high attention of researchers.Inspired by such verification of concepts,we designed the Photoinduced Electron Transfebetween DNA-AgNCs and the G-quadplex and hemin.Based on this novel DNA-AgNCs and the G-quadplex and hemin PET system,a molecular beacon-like model was developed to detect mRNA.Based on the PET possess of is novel DNA-AgNCs and the G-quadplex and hemin the fluorensence biosensor towards target bio molecules especially the DNA and RNA was realized.The detection limit for mRNA detection is as low as 1.8 nM.Based on this strategy,low background interference and high sensitivity were realized.