The Synthesis And Bioimaging Application Of N-doped Carbon Quantum Dots

With the rapid development of nanotechnology,carbon quantum dots(Cdots),as novel fluorescent nanoparticles,have many excellent fluorescence properties.These include high fluorescence quantum yield,high resistance to photobleaching,up-conversion emission characteristics,long fluorescence lifetiome,which are comparable to organic fluorescent dyes and semiconductor quantum dots.The Cdots have been widely developed and applied in the fields of biosensing,targeted tracing,drug delivery and optoelectronic devices.In terms of bioimaging,the vast majority of unmodified carbon dots are not specific for cancer cells and lack the ability to target recognition.Moreover,in vitro fluorescent labeling has only been applied to two-dimensional adherent cells,which have many limitations compared to solid tumors in vivo.Therefore,Cdots with high fluorescence properties and biological affinity were prepared in this thesis,and the photostability and long-term tracing ability of the nanoprobes in cells were studied.A fluorescent labeling model for three-dimensional multicellular tumor spheroids was established.The visual recognition and detection methods for hepatocellular carcinoma cells and gastric cancer cells were realized by different fluorescence distributions and strong fluorescence contrasts.The main contents are as follows:(1)The carbon quantum dots(o-Cdots)using o-phenylenediamine(oPD)as precursor were synthesize through one-step hydrothermal synthesis,which possessed particle size of 4 nm,good dispersibility,excellent and stable fluorescence properties,abundant surface functional groups and high biocompatibility.(2)Through fluorescence tracing of several cell generations,we applied the Cdots to cell imaging and explored their long-term imaging ability in cells.The experimental results show that the Cdots entered the cells and distributed in the cytoplasm and nucleus,and exhibited stable fluorescence labeling intensity.Moreover,compared with commercial nucleus dyes(DAPI),the Cdots didn't present obvious photobleaching after 30 minutes of continuous ultraviolet irradiation.In addition,the strong fluorescence from the Cdots could still be clearly observed after five generations.A three-dimensional cell culture model was established.The natural porous scaffolds and matrix materials were used to simulate the extracellular matrix in the micro-environment of tumors.Multicellular cancer spheroids cultured from gastric cancer cells were labeled with the above carbon dots.Laser confocal microscopy was used to observe the fluorescence distribution and imaging depth of Cdots.Experiments showed that Cdots could label the whole tumor spheroids within 24 hours.These superior properties make Cdots develop an ideal non-invasive fluorescent probe for longterm tracking imaging.(3)The dependence of fluorescent labeling sites of Cdots on cell types was investigated.The Cdots without any targeting modification could label different locations of hepatoma cells(HepG2)and normal hepatocytes(LO2).For HepG2 cells,the Cdots could rapidly permeate the cell membrane and diffuse into cytoplasm and nucleus within 3 minutes,and hold their location in the targets for 24 h.However,the Cdots exhibited bright fluorescence only in cytoplasm of LO2 cell lines.In addition,using hyaluronic acid(HA)as a target molecule to modify carbon dots,the active targeting labeling of CD44 overexpressed gastric cancer cells was realized.Compared with normal gastric cells,the recognition and detection of gastric cancer cells were realized with obvious fluorescence contrast.Therefore,the Cdots have great potential for rapid and selective bioimaging applications,and are expected to be used in the screening of cancer cells.