Gold Nanoparticale Involved Surface Energy Transfer System For Antibiotic Detection And Tumor Nucleic Acid Imaging

Surface energy transfer(SET),also known as nanometal surface energy transfer(NSET),is one of the sensitive optical analysis techniques of resonance energy transfer,which is widely used in biomolecular structure and knetic characterization,biological imaging,analysis and detection,disease diagnosis and treatment,etc.NSET is the process of transfer energy from donor fluorophore transferring energy to nanometal surface(gold nanoparticle)as energy acceptor in a non-radiative manner.Compared with traditional F?rster resonance energy transfer(FRET),NSET has a higher energy transfer efficiency and a longer effective distance.The energy donor-acceptor pairs are essential in energy transfer systems.Common energy donor and acceptor include organic small molecules(organic dye molecules and small molecule quenchers)and nanomaterials.Among them,organic small molecules have the advantages of simple structure,easy labeling and wide spectral range,which are used as energy donor and acceptor.Nanomaterials are favored by researchers from all walks of life because of their unique properties.Especially gold nanoparticles(AuNPs)in precious metal nanomaterials have broad development prospects in catalysis,sensing and analysis due to excellent properties such as simple synthesis and easy modification,good stability and biocompatibility.AuNPs are widely used as energy acceptors in surface energy transfer systems because of a high molar absorption coefficient,hence the prepared fluorescent probes based on NSET strategy have been continuously developed in the field of analytical sensing.At present,most of the surface energy transfer systems based on AuNPs used the nucleic acid labeling strategy to functionalize dense energy donors on the surface of AuNPs.The use of AuNPs as a single energy acceptor caused problems such as high background signal and low sensitivity.The functionalization process involved multiple steps of salt addition and centrifugation,making the preparation process cumbersome and time-consuming.In order to solve these problems,another energy acceptor or electron acceptor was modified on the surface of AuNPs to improve the fluorescence quenching performance,and the constructed dual energy transfer nanoprobe and energy transfer-electron transfer coupled fluorescent nanoprobe were used for antibiotic detection and tumor nucleic acid imaging analysis.The specific two parts of the research are as follows:(1)Preparation of “enhanced nanoflares” and sensitive detection of ampicillin.The novel enhanced nanoflare was prepared by introducing the black hole quencher(BHQ-2)into the conventional spherical nucleic acid(SNA)nanostructure and used for ampicillin analysis.First,the ampicillin aptamer partially hybridized with the TAMRA-labeled short-chain DNA(TAMRA-cDNA)to form flares.Then,the Au-S bond was used to modify the flares to the surface of AuNPs to obtain the traditional nanoflares.Finally,in order to prepare enhanced nanoflares,BHQ-2 was functionalized on the surface of AuNPs via Au-N bond.Due to the close distance between the donor and acceptor,the occurrence of NSET and FRET,and the fluorescence of the donor TAMRA was quenched by AuNPs and BHQ-2 synergistically.When ampicillin was added to the enhanced nanoflares probe,ampicillin specifically bound to the aptamer and triggered the fluorescence recovery of TAMRA in a concentration-dependent manner.Compared with traditional nanoflares,the dual-receptor strategy greatly reduced the background signal and improved the energy transfer efficiency between the donor and acceptor,thereby achieving sensitive detection of ampicillin.The detection limit of ampicillin was 0.65 ng/m L and the linear range was 1.8-20 ng/m L.It has been successfully used for the detection of ampicillin in spiked milk samples and pharmaceuticals.(2)Preparation of energy transfer-electron transfer coupled fluorescent nanoprobe and mRNA imaging in living tumor cells.First,taking advantage of the self-polymerization properties of dopamine(DA)under alkaline conditions,polydopamine(PDA)-coated AuNPs(Au @ PDA NPs)were prepared.Then,using the characteristics of PDA adsorbing single-stranded DNA(ssDNA)without interacting with double-stranded DNA(dsDNA),fluorescent nanoprobe coupling energy transfer and electron transfer based on simple physical adsorption dye labeling ssDNA(dye-ssDNA)in one step.PDA is a good electron acceptor due to its oxidized quinone structure,and AuNPs as energy acceptor.Due to the close distance between the donor and acceptor,the quenching efficiency of the donor Cy5 was as high as 90% via NSET and photoinduced electron transfer(PET).In presence of target tumor-related mRNA,dye-ssDNA and mRNA were completely complementary to form ds DNA,then releasing from the surface of Au @ PDA NPs,and the fluorescence signal of Cy5 was greatly recovered,enabling sensitive analysis of mRNAs in solution.The detection limit of TK1 mRNA was 0.43 n M,and the linear range was 1.8-90 nM.Au @ PDA NPs as highly efficient carrier and quencher have excellent biocompatibility.Nanoprobe entered cells through endocytosis,thereby enabling imaging of tumor-related mRNAs at the cellular level simultaneously.In summary,in this study,we selected suitable donor and acceptor to functionalize AuNPs and constructed biosensing system based on the surface energy transfer,which improved the energy transfer efficiency and made the low background of nanoprobes to improve the sensitivity and accuracy of the detection.At the same time,nanoprobes were applied to sensitive detection of antibiotic and in situ imaging of tumor nucleic acids at the cellular level.