The Study Of Developing Novel Biosensors Based On Noble Metal Nanoparticles

Noble metal nanoparticles,especially gold nanoparticle and silver nanoparticle,can have many advantageous potential applications in biosensing,owing to their excellent biocompatibility,prominent electrical and optical properties,as well as the particle surface ameniable to chemical modifications.Moreover,noble metal nanoparticles can also interact with various types of targeting probes such as nucleic acids and peptides,while diverse tool enzymes of these probes enable a great many of signal amplification protocols,which provides a proper opportunity for noble metal nanoparticles in biosensing application.In this thesis,noble metal particles are utilized to transfer the recognition of nucleic acid or peptide probe with the analytes to optical or electric signal.In the meanwhile,the chem-physical properties of the nanoparticles are employed to enhance the effect of signal readout,thus some ingenious signal amplification steps initiated and/or controlled by nucleic acid-and peptide-modified nanoparticles are designed to further strengthen the amplification efficiency,enabling sensitive detection of various types of analytes related to human health.Taking advantage of the properties of noble mental nanoparticles combined with the nucleic acid-or peptide-based reactions for signal generation and amplification manipulated by the nanoparticles,several new methods have been proposed for the analysis of proteins,nucleic acids and metal ions which are related with the health of human beings.The details are as follows:1.Assay of Streptavidin Based on PLA-dependent Ribozyme Cleavage Cycling Controlled Aggregation of Gold NanoparticlesIn this work,we describe a gold nanoparticle(AuNP)colorimetric biosensor for streptavidin based on PLA-dependent DNAzyme.A target-initiated proximity ligation assay(PLA)protocol with DNAzyme formation is for the first time designed for ultrasensitive colorimetric detection of protein.This protocol ingeniously converts the recognition of target protein with multiple ligands to assembly of DNAzyme,greatly improving the selectivity and sensitivity of detection.What's more,DNAzyme cyclically catalyzes cleaving substrate to amplify the readout signal without other protease,simplifying the reaction system and improving the stability.Further,the whole reaction only needs two steps in the same solution,without any requirement for additional cleaning process.Besides,the results are visible by naked eye,so this biosensor has potential for point-of-care analysis.Furthermore,this biosensor can be a universal platform to assay other target with multiple ligands by changing the ligands.2.Assay of Gamma-glutamyltranspeptidase Based on in situ Reduction and Anodic Striping of Silver Nanoparticles Mediated by Graphene OxideIn this work,we have fabricated a novel gamma-glutamyltransferase(?-GGT)sensor based on in situ reduction and the solid-state voltammetry of silver nanoparticle(AgNP)mediated by graphene oxide(GO).Specifically,glutathione(GSH)is firstly immobilized on the surface of a gold electrode,and GO nanosheet itself is introduced to recognize the charge change of GSH caused by the catalysis of-GGT.Then,the in situ reduction of AgNP is formed on the GO surface,in which only 10 minutes would be needed and a large number of AgNP with homogeneous dimension could be produced.Since the product may have excellent electric conductivity and the solid-state voltammetry of Ag/AgCl can provide a well-defined symmetrically sharp silver stripping peak,a sensitive electrochemical biosensor for the detection of y-GGT is thus developed.Moreover,the experimental results obtained in this work indicate that AgNP of in situ reduction can be a promising material in a general sensing platform and GO itself can serve as a general recognition element for biosensor fabrication,so such kind of more biosensors can be developed in the future.3.Assay of Trypsin Based on Peptide Probe-functionalized Silver NanoparticlesHerein,we report a simple,sensitive,label-free colorimetric assay of trypsin based on AgNP aggregation.Generally,a specially designed short peptide chain acts as both the stabilizer of AgNP and the substrate of trypsin.In the presence of trypsin,the negatively charged part of peptides will be hydrolyzed,leaving the positively charged dipeptide capped on the surface of AgNP.The electrostatic property alteration then leads to the AgNP's aggregation in certain salt condition.The solution color may change correspondingly due to the localized surface plasmon resonance,which can be monitored by naked eye and UV-vis spectrophotometry.This novel AgNP-based colorimetric method for quantitative determination of trypsin has a linear detection range from 2.5 to 200 ng/mL and a rather low detection limit down to 2 ng/mL.The determination of trypsin can also be realized in complex biological fluids by the proposed method,demonstrating its great potential utility in the clinical applications in the future.4.Detection of MicroRNA Based on Anodic Striping of Silver Nanoparticle and Strand Displacement ReactionMicroRNAs are not only important regulators of a wide range of cellular processes,but also identified as promising disease biomarkers.Due to the low contents in serum,microRNAs are always difficult to be accurately detected.Herein,an electrochemical biosensor for ultrasensitive detection of microRNA based on anodic striping of AgNP and strand displacement amplification is developed.The stand displacement amplification mediated by DNA on the AgNP and anodic striping of AgNP greatly improve the sensitivity of microRNA biosensor.Beside,four DNA single strands are engineered to form a tetrahedral nanostructure with a pendant stem-loop,and modified on a gold electrode surface,which largely enhances the molecular recognition efficiency.Moreover,ultra-high sensitivity with the limit of detection(LOD)down to 0.4 fM and the satisfactory results in the real samples analysis conform this biosensor great clinical value,which may have practical utility in early diagnosis and prognosis of certain diseases.5.Detection of Silver Ion Based on Dual Signal Amplification by Gold Nanoparticle and Enzyme Cleavage ReactionSilver ion(Ag+)is a highly toxic heavy metal ion to fungi,viruses,bacteria,and animals.Therefore,Ag+monitoring in water or food resources has become-extraordinarily important within the scope of human health.Here,we report a AuNP and enzyme cleavage based dual signal amplification strategy for ultrasensitive detection of Ag+using electrochemical techniques.This sensing platform for Ag+has an extremely low detection limit of 470 fM,which also has satisfactory selectivity.Besides,it can be directly used in drinking water and lake water samples with stable results.Thus,this biosensor has great potential in the environmental monitoring and testing.Moreover,the strategy proposed in this work can be further developed as a generalized platform for the detection of other analytes by designing new DNA sequences for specific recognition.