| Objective: 1.Water-soluble Cd Te quantum dots(QDs)with strong fluorescence signal,excellent chemical stability,good crystallinity,small size and uniform dispersion were prepared by simple and cheap hydrothermal method.2.The QD-PEG-Ab-AFP probes with strong fluorescence signal and high sensitivity for hepatocellular carcinoma cells was constructed by chemical synthesis method.3.The QD-PEG-Ab-AFP probes was used to Label liver cancer PLC /PRF /5 cells..Methods: 1.The preparation of quantum dots and characterization of QDs crystal structure,morphology and optical properties In this study,water-soluble Cd Te quantum dots(QDs)were synthesized by hydrothermal method in the aqueous phase.X-ray diffraction,Transmission electron microscopy(TEM),Dynamic light scatterometer(DLS),were used to observe the crystal structure and morphology of QDs.The optical properties of QDs were tested with Ultraviolet-Visible(UV-Vis)spectrophotometer and Fluorescence spectrophotometer.2.The construction of QD-PEG-Ab-AFP probes PEG and QDs were combined through chemical synthesis method.PEG bioconjugation can not only improve the biocompatibility of QDs,but also provide the conditions for coupling alpha-fetoprotein antibody(Ab-AFP).Then the QD-PEG-AbAFP probe was fabricated by coupling the monoclonal antibody Ab-AFP to the surface of QD by the chemical route.The effect of AFP coupling on QDs was observed by SDS-PAGE.Transmission electron microscopy(TEM),Dynamic scatterometer(DLS),UV-Vis spectrophotometer and fluorescence spectrophotometer were examined to observe the morphology and optical properties of QD-PEG-Ab-AFP probes respectively.3.The QDs probes labeling liver cancer PLC / PRF / 5 cells The hepatocellular carcinoma cells were labeled with QD-PEG-Ab-AFP probe by quantum dots-based immunofluorescence labeling,and the labeling results were observed and photographed under confocal laser scanning microscope.Results: 1.The crystal structure,morphology and optical properties of QDs(1)A series of QDs solution with different colors were successfully prepared by controlling the hydrothermal conditions.The color of the QDs was light,yellow,red and magenta in natural light.While they showed pale green,green,yellow,red and other color fluorescence under the 365 nm ultraviolet light.Moreover,QDs could disperse uniformly in natural environment at room temperature.(2)QDs were mainly composed of Cd and Te elements,which were cubic sphalerite structures.The particles were uniform spherical shape whose size is about 4 nm.(3)There was a shoulder at 540 nm with a smaller peak width in the UV-Vis Absorption Spectra of QDs.The fluorescence spectra of QDs had a narrower and symmetrical peak at about 590 nm,which indicated that the particle size distribution was relatively narrower.And the fluorescence of QDs did not occur perceive change even if the QDs solution was kept over 10 months.2.Construction of QD-PEG-Ab-AFP probes(1)In SDS-PSGE,there were red fluorescence band and Coomassie blue band at the same position of the film,so it was proved that QD and Ab-AFP were coupled together successfully.(2)The size of QD-PEG-Ab-AFP probe modified with Ab-AFP was 5 nm,which increased about 1 nm comparing with QD.DLS results showed that the hydrodynamic diameter of the QD-PEG-Ab-AFP probe increased from 10 nm to about 95 nm compared with that of QD.However,The QD-PEG-Ab-AFP probe modified with AbAFP is still spherical,dispersed evenly and has similar particle size.(3)The UV-Vis absorption spectrum of the QD-PEG-Ab-AFP probe was a continuous curve and has a shoulder at 540 nm.The absorption curve of QD-PEG-Ab-AFP probe was similar to that of QD,which indicated that the coupling of Ab-AFP does not change the structure and absorption properties of QD.However,the absorption intensity of QD-PEG-Ab-AFP probe reduced slightly.The fluorescence spectrum of the QD-PEGAb-AFP probe was also a continuous curve and had a narrow and symmetrical emission peak at 585 nm.Compared with the fluorescence spectra of QD,the shape of fluorescence spectrum of QD-PEG-Ab-AFP probe did not change and the fluorescence intensity did not decrease,but the peak possession of QD-PEG-Ab-AFP probe was blue-shift by 5 nm.3.Labeling for liver cancer PLC / PRF / 5 cells No fluorescence signal could be found in liver cancer cells labeled by QD-PEGs,which indicated that the modification of PEG decreased and even eliminated the nonspecific adsorption of QDs.In contrast,the strong red signals could be observed in cytoplasm in HCC cells labeled by QD-PEG-Ab-AFP probes,which revealed that the QD probes could specifically and accurately attach to AFP antigen and keep stable optical properties.Conclusions: 1.In this study,a series of water-soluble Cd Te-QD with different colors were successfully prepared by hydrothermal method.The QDs were cubic sphalerite structure.The particles were uniform spherical shape with diameter about 4 nm.They performed good dispersity,long fluorescence stability that they could be stored for at least 10 months,which demonstrated that QDs has excellent chemical stability and fluorescence stability.2.The QD-PEG-Ab-AFP probe was successfully obtained by coupling the AFP antibody to the surface of the QD by chemical approach.The size of the QD-PEG-AbAFP probe was increased to about 5 nm.Although the size of QD-PEG-Ab-AFP probe increased because of the coupling of AFP antibody,it did not change the QD structure.The fluorescence property of QDs did not change significantly.The absorption spectra and fluorescence spectra of the QD-PEG-Ab-AFP probe were similar to those of the QD,indicating that the coupling of AFP with QD did not change the structure and properties of QD.3.The synthesized QD-PEG-Ab-AFP probe could accurately label Liver cancer PLC/PRF/5 cells with strong fluorescence signal and excellent fluorescence stability. |
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