Synthesis Of Silver And Gold Nanoclusters For Biological And Chemical Analysis

Noble metal nanoclusters, especially silver and gold nanoclusters (AgNCs and AuNCs), can exhibit molecule-like electronic transitions between HOMO-LUMO energy level due to their small size. Owning to their unique electronic structure as well as physical and chemical properties, the noble metal nanoclusters have attracted much attention recently and have been widely used in biosensing, biolabeling, optical imaging, single-molecule imaging, and so on. Compared to organic fluorescent dyes, semiconductor quantum dots, and fluorescent polymer nanoparticles, the noble metal nanoclusters have many advantages including non-toxicity, non-photobleaching, and tunable emission. However, there are still some shortcomings related to the synthesis and applications of noble metal nanoclusters such as complicated preparation process, low fluorescent quantum yield, and poor stability. To address the above issues, we have systematically studied the synthesis of AgNCs and AuNCs as well as their applications in fluorescent analysis and imaging. This thesis includes the following two parts:Part Ⅰ Synthesis of AgNCs for biological and chemical analysis, including the following two parts:Based on the selective recognition of cytosine and Ag+, we designed a specific oligonucleotide as the stabilizer and achieved the synthesis of AgNCs with strong fluorescence and excellent biocompatibility. The used oligonucleotide contains half of cytosine bases and half of adenine bases, which is not easy to be self-folding or hybridization. The as-prepared AgNCs were further used to detect thiol-containing drugs. The results demonstrated that thiol-containing drugs could selectively interact with AgNCs via the strong Ag-S interaction, and effectively quench the fluorescence of AgNCs following a simple static quenching mechanism. A new spectrofluorometry for highly selective and sensitive detection of captopril was proposed, showing that the present method has the advantages of speediness, accuracy, high selectivity and sensitivity for the assay. In order to confirm the practicality of this method, it has been used for the determination of captopril in tablets.Then, a facile strategy for highly selective and sensitive detection of bacterial alarmone, ppGpp, which is generated when bacteria face stress circumstances such as nutritional deprivation, has been established. Herein, based on the strong fluorescent quenching effect of Cu2+on DNA-AgNCs as well as the specific recognition between Cu2+and ppGpp, we report a "turn-on" fluorescent method for the detection of ppGpp.This work not only achieved highly selective detection of ppGpp in a broad range concentration of2-200μmol/L, but also improved our understanding of the specific recognitions among DNA-AgNCs, Cu2+, and ppGpp. This strategy has also identified that AgNCs functionalized with a specific molecule on their surfaces can be engineered as a novel fluorescent probe for a wide range of applications such as biosensing and bioimaging.Part Ⅱ Synthesis of AuNCs for optical imaging, including the following three parts:Firstly, we rapidly synthetized bovine serum albumin (BSA) stabilized Au20nanoclusters (BSA-Au20NCs) with high fluorescence quantum yield (QY) up to15%. The success of this synthesis relied on the rational manipulation of reaction kinetics and screening of an appropriate ligand to stabilize the formed AuNCs. This new protocol for the synthesis avoided complex treatment, long-time preparation process, and the use of toxic solvents. More importantly, the as-obtained Au20NCs have a lot of advantages including small sizes, high fluorescence QY, excellent photostability, non-toxicity, and good stability in biological systems, which make them ideal candidates for optical imaging in vitro and in vivo.Except for solution phase, the AuNCs can also be synhesized on the surfaces of solid substrates. We demonstrated that luminescent silk and fabric could be produced through nanotechnology-in situ chemically coating with luminescent AuNCs on the surface of natural silk fiber. This syntheis relied on a redox reaction between protein-based silk and Au salt precursor. The luminescent silk coated with AuNCs (we call it as golden silk) possesses good optical properties including relatively long wavelength emission, high quantum yield, long fluorescent lifetime, and photostability. Moreover, the as-prepared golden silk showed better mechanical property (1.3times) and performance to interdict UV light (1.5times) than pristine silk, together with good biosafety. Benefiting from these advantages, as a concept of proof, we further demonstrated that such photoluminescent silk was an excellent candidate for anti-counterfeiting. This work not only provided an effective strategy for in situ preparation of luminescent metal nanoclusters on solid substrate, but also paved the way for large-scale fabrication of novel silk-based materials or fabrics through nanotechnology.Finally, we prepared a functionalized Au20NCs probe for the recoginition of tumor sites via active-targeted mode. The previous studies on the uptake of nanoprobes for imaging are usually based on the passive mode (e.g., endocytosis). In this case, the efficiency for the uptake would be relatively low, and the probes were also not able to selectively accumulate at tumor sites. To solve the above issues, our work indicated that the uptake of Au20NCs by both cancer cells and tumor-bearing nude mice could be improved by receptor-mediated internalization, compared with that by passive targeting. The results demonstrated that the functionalized-Au20NCs were excellent probes for active tumor-targeted imaging in vitro and in vivo. We believe this general strategy of active tumor-targeted optical imaging based on the functionalized Au20NCs is promising can be applied in clinical cancer diagnosis and therapy in the future.In summary, we have synthesized biomolecule-stabilized AgNCs and AuNCs with excellent fluorescenct properties and further used them for a variety of purposes. On the one hand, we have designed oligonucleotide-stabilized AgNCs and successfully applied them in drug assay and the detection of important signaling molecule in bacteria. On the other hand, we have prepared BSA-stabilized AuNCs in solution phase and on solid substrate, respectively. By manipulating the reaction kinetics, AuNCs with high fluorescent QYs and stability can be obtained. Then, we employed the functionalized-AuNCs as optical nanoprobes for active tumor-targeted imaging in vitro and in vivo. The established methods for the synthesis and applications of noble metal nanoclusters were simple and effective, and thus could be expanded to other ordinary laboratories. The innovation of this thesis is mainly in the following two aspects:firstly, we developed simple approaches for the synthesis of noble metal nanoclusters with good properties; secondly, we developed effective platforms for biosensing and bioimaging based on the excellent fluorescent properties of noble metal nanoclusters.