The Construction And Application Of Fluorescent Biosensors Based On Carbon Nanomaterials And Hybridiation Chain Reaction

In recent years,fluorescent biosensors have been used in environmental detection,disease diagnosis and prognosis,biological drug delivery and other fields based on their advantages,such as simple operation,fast signal response and good selectivity.In addition,as a new type of nanomaterials,carbon nanomaterials have been widely applied in many fields such as electronic devices,optical components,energy development,medical drug delivery and so on.As one of the typical representatives,carbon dots have attracted extensive attention due to their low toxicity,strong photobleaching resistance and good biocompatibility,and many research has been developed.However,due to the low content of many substances to be measured and the interference of the surrounding environment,the fluorescence signal amplification strategy is always used to improve the sensitivity,accuracy and timeliness of the detection of ions,DNA,protein and other substances in vivo.Hybridization chain reaction?HCR?has become an efficient molecular amplification tool due to its enzymatic-free,high isothermal amplification efficiency,good sensitivity and structural flexibility.It has broad application prospects in biosensors construction,medical imaging,diagnosis and treatment.At present,HCR has been applied in the detection of various targets,such as DNA or RNA,proteins,small biological molecules and metal ions.Combining the unique advantages of carbon nanomaterials with hybridization chain reaction,a series of fluorescence-based biosensors were constructed for Cu²⁺ biosensor and cell imaging,as well as the detection of circulating tumor DNA and silver ions,and applied them into the analysis of actual sample systems.Specific research contents are as follows:?1?A new type of high fluorescent sulfur,nitrogen and oxygen co-doped carbon dots?S,N,O-CDs?was synthesized by hydrothermal method using m-phenylenediamine and sulfonamide as raw materials.The results show that the prepared S,N,O-CDs have high doping ratio and fluorescence yield.It has good fluorescence stability,excellent selectivity towards Cu²⁺ and good linear fluorescence response in the range of 2-60 ?M.The detection limit is 0.29 ?M.Using the advantages of S,N,O-CDs,such as good selectivity and low cytotoxicity,it was applied to Cu²⁺ biosensor and imaging in cells.In addition,based on the strong complexation of Cu²⁺ with pyrophosphate ion?PPi?and the specific hydrolysis ability of alkaline phosphatase?ALP?to PPi,the quantitative analysis of PPi and ALP was further realized by S,N,O-CDs /Cu²⁺ system.Moreover,the method also realized the detection of PPi and ALP in real samples.?2?Based on hybridization chain reaction?HCR?and graphene oxide?GO?,a label-free and enzyme-free fluorescence detection platform was constructed to detect circulating tumor DNA?ct DNA?.According to the sequence of ct DNA,two related hairpin structures were designed by consulting literatures.Based on the special binding effect of SYBR Green I fluorescent nucleic acid dye?SG?on double-stranded DNA,when the target ct DNA does not exist,the hairpin structure can be stably existed in the reaction system separatly,and stacked on the surface of graphene oxide through the p-p action.With the addition of target,hybridization with hairpin H1 occurred,which further triggered HCR and resulted in long double-chain products far away from the surface of graphene oxide,and the fluorescence signal of the system was enhanced.Quantitative analysis of ct DNA was achieved with the change of the fluorescence signal.?3?A fluorescence-enhanced biosensor based on graphene oxide and hybridization chain amplification technology is constructed for the analysis and detection of silver ions.Firstly,the auxiliary probe rich in cytosine C and two kinds of related stem-ring structures were designed through literature review.Under the action of SG dyes,when Ag + does not exist,free nucleic acid probes adsorb on GO surface by the interaction of p-p action,resulting in the decrease of the fluorescence signal.With the addition of Ag+,the specific interaction between Ag+ and cytosine C will cause the formation of C-Ag+-C structure,which leads to the HCR occurred.The obtained HCR products were far away from the surface of graphene oxide and produced strong fluorescence signals,and finally the quantitative detection of Ag+ was realized.