Synthesis Of Functional Near-Infrared Ag₂S Quantum Dots And Their Applications In Biosensing And Bioimaging

Compared to visible light quantum dots,near-infrared quantum dots?NIR QDs?have good tissue penetrability,which can avoid the influence of weak penetration and organism autofluorescence,so they have attracted extensive attention in biological imaging and disease diagnosis.The novel Ag₂S quantum dots?Ag2S QDs?have attracted great research interest for their excellent NIR fluorescence properties and biocompatibility.At present,most researches of Ag₂S QDs focus on preparation methods and in the fields of photodetectors,solar cells,and catalysis.And in the fields of in vivo imaging,photothermal therapy,and tumor targeting.In terms of the synthesis and functionalization of Ag₂S QDs,the superior synthesis and functionalization methods to improve their quantum yield and enhance stability is still challenging.About the analysis and application of Ag₂S QDs,there is a broad space for imaging analysis and detection of small molecular or cancer markers in vivo based its superior NIR fluorescence properties.Therefore,the strategy of synthesis and functionalization and the bioanalytical applications of NIR Ag₂S QDs in-depth and systematic research are of great significance.In this paper,we focus on the preparation and functionalization of NIR Ag₂S QDs,as well as the construction of corresponding fluorescence analysis systems,for bioimaging and cancer marker detection.The main contents are as follows:Chapter 1.IntroductionThis chapter mainly introduces the nature,types and advantages of NIR QDs.And the properties,preparation methods and applications of Ag₂S QDs were systematically summarized and explained.Additionally,it briefly introduces the work content and significance of this paper.Chapter 2.One-step synthesis of mercaptopropionic acid functionalized hydrophilic NIR Ag₂S QDs in aqueous phaseNIR QDs have been widely used in biologically relevant fields due to their unique optical properties of deep penetrating ability and weak autofluorescence in living organisms.However,the toxicity caused by the synthesis of raw materials and organic phases limits its application.Therefore,this chapter mainly explores the use of simple,environmentally friendly methods to prepare NIR Ag₂S QDs with low toxicity and high stability.We have found that 3-mercaptopropionic acid can be used as a stabilizer to synthesize good performance of NIR Ag₂S QDs in the aqueous phase.And,the method is simple and environmentally friendly.In the experiment,Ag₂S QD with good dispersibility and uniform particle size?2.49 ± 0.64 nm?and shows a tunable photoluminescence emission?732-801 nm?in the first near-infrared?NIR-I?region by simply adjusting the composition ratio of the raw materials,the temperature and time of the reaction system,and the p H value.The experimental results also show that the quantum efficiency of Ag₂S QDs is up to 14%?ICG=13%,DMSO?.And the Ag₂S QDs have good light/colloid stability and ultra-low cytotoxicity.These properties indicated this Ag₂S QDs expected to be effective nanoprobes for NIR biological window bioimaging.Chapter 3.Rare earth ions enhanced NIR Ag₂S QDs for the detection of fluoride ions and cell imagingIn this chapter,a novel phenomenon was discovered that the fluorescence intensity of Ag₂S QDs could be enhanced in the presence of rare earth ions through aggregation between-COOH and rare earth ions.Meanwhile,a “turn-off” fluoride ion?F-?fluorescent nanoprobe was constructed based on the strong soft-hard acid-base binding ability between F-and rare earth ion.Interestingly,the fluorescence intensity of QDs was obviously enhanced upon the addition of various rare earth ions,especially in the presence of Gd³⁺.The AIE mechanism was proved via the TEM,Zeta potential and dynamic light scattering analysis.Moreover,the coordination between rare earth ions and F-could lead to the quenching of fluorescence QDs due to the weakening the AIE.Based on these findings,we developed a highly sensitive and selective method for detection of F-with a linear range of 5-260 ?M and a detection limit of 1.5 ?M.Moreover,it was proved by cytotoxicity experiments that the prepared Gd³⁺-Ag₂S QDs also have good biocompatibility.Combined with the unique advantages of Ag₂S QDs in the NIR region,the label-free NIR fluorescence probe was successfully used for Fbioimaging in live cells.Chapter 4.LRET system based on upconversion luminescence nanoprobe-Ag₂S QDs for ratio sensing and imaging of p H in vivoIn this chapter,the obtained MPA-Ag₂S QDs were further modified by small molecule GSH to obtain GSH and MPA co-modified Ag₂S QDs?GM-Ag₂S QDs?probe with excellent p H response,which based on ligand exchange.The upconversion nanomaterials?UCNPs?were coated with nano-silica assembly with Ag₂S QDs was achieved by coupling reaction.And by adjusting the thickness of the nano-silica layer,a luminescence resonance energy transfer?LRET?system was successfully developed using NIR Ag₂S QDs as the energy acceptors and UCNPs as the energy donors.The system with the feature of NIR excitation?980 nm?and NIR emission?795 nm?.Based on the p H response of GM-Ag₂S QDs and the optical properties of UCNPs,a ratiometric p H fluorescent probe was constructed and successfully applied to intracellular p H sensing with a response range of 5.0 to 9.0.Based on the NIR excitation-NIR emission and the high-resolution advantages of ratiometric luminescence nanoprobes in bioimaging,we successfully applied this nanoprobe to the differentiation of tumors and normal tissues in the zebrafish model.Chaper 5.Assembly of aptamer functionalized Ag₂S QDs and magnetic nanoparticles for specific recognition and high sensitivity detection CTCsWe further utilized the aptamer sequence to functionalize the NIR Ag₂S QDs for ultrasensitive fluorescent labeling of circulating tumor cells?CTCs?.Firstly,DNA1 functionalized Ag₂S QDs?DNA1-Ag2S?were synthesized by one-step method based on the phosphorothioated DNA sequence.The hairpin structures H1 and H2 were then separately designed for assembly with DNA1-Ag₂S and aptamers?specific binding to MUC1 overexpressed on the surface of MCF-7 cells?.And then,the NIR fluorescent Ag₂S nanoassembly successfully constructed through hybridization chain reactions using aptamer-modified Ag₂S QDs,which can extremely improve the imaging sensitivity and reduce background signal of blood samples.Moreover,due to the NIR characteristics and multi-branched structure of the probe,not only can the sensitivity of cell imaging be improved,but also the binding ability of the multi-aptamer structure to tumor cells is stronger than that of a probe having a single aptamer.In addition,the antiepithelial-cell-adhesion-molecule-antibody?Anti-Ep CAM?labeled magnetic nanospheres have been used for highly capture rare tumor cells in whole blood.That is,NIR fluorescent Ag₂S QDs based signal amplification combing with immune-magnetic spheres?IMNs?for highly efficient magnetic capture and ultrasensitive fluorescence labeling of CTCs.The probe can detect at least 6 cancer cells in clinical samples.Chaper 6.Assembly of composite cell membrane encapsulated magnetic beads and multi-aptamer functionalized Ag₂S QDs for specific recognition and ultrasensitive detection CTCsSince CTCs are extremely low in peripheral blood and susceptible to white blood cell interference,it is still challenging to efficiently capture and detect CTCs.In this chapter,we further designed a mutil-aptamer-modified Ag₂S QDs?Tetra-DNA-Ag₂S QDs?,and encapsulated the magnetic nanoparticles Fe3O4@Si O2 with tumor cells membranes?TM?and white blood cells membranes?WM?to obtained the WT-HM-Fe3O4@Si O2.WT-HM-Fe3O4@Si O2 further modified with SA?SA-WT-HM-Fe3O4@Si O2?and bingding with Tetra-DNA-Ag₂S QDs modified with biotin to significantly increasing the sensitivity,reliability,and specificity of the cell capture.The probe has the following advantages:?1?NIR Ag₂S QDs modified by Tetra-DNA aptamers can effectively improve the targeting recognition efficiency of CTCs;?2?can effectively avoid white blood cells in the sample by encapsulation of WBC homology interference;?3?modification of tumor cell membranes can increase the targeting of CTCs.Based on the above characteristics,the integrated probe can effectively improve the capture efficiency and analytical sensitivity of CTCs.In the experiment,it was found that the Tetra-DNA-Ag₂S QDs can increase the capture efficiency of CTCs from 62.26% to 97.20% compared with the single-aptamer modification.And,compared with the probe without cell membranes,the capture efficiency of the probe for CTCs increased from 73.02% to 97.63%.