Preparation Of Fluorescent And Porous Colloidal Particles And Their Interactions With Cells

Along with the fast development of nanotechnology and nanomedicine,the colloidal particles are usually integrated with multifunction such as tailored wettability,specific targeting,imaging and controlled release of the desired substances to be used as carriers for drug control release,cancer and gene therapy,disease diagnosis and bio-imaging via the way of intracellular uptake and delivery.First,we synthesized the water-soluble 3-mercaptopropyl acid(MPA) capped CdTe nanocrystals and optimized the preparation conditions.Results showed that when the pH of precursor was fixed at 9.0 and the molar ratio of Cd:Te:MPA is 1:0.2: 2.4 with a Cd²⁺ concentration of 1.25mM,the high quality CdTe nanocrystals could be obtained under a high pressure and temperature condition with a quantum yield up to more than 40%.The QDs could also be incorporated into inorganic colloidal particles.The MPA-CdTe quantum dots(QDs) were embedded into CaCO₃ microparticles with a size of 1.4～4.4μm by addition of the QDs into Ca(NO₃)₂ solution during a mineralization process.Compared to the parent QDs,about 1/7～/1/4 photoluminescence efficiency of the embedded QDs was preserved,enabling the CaCO₃(CdTe) particles visible under UV irradiation.The structure and morphology of the CaCO₃(CdTe) particles were characterized by X-ray diffraction(XRD),UV-vis spectroscopy,scanning electron microscopy(SEM),transmission electron microscopy (TEM) and confocal laser scanning microscopy(CLSM).Protected by the CaCO₃ particles,the QDs in the composites were more stable against long term storage,UV irradiation and cell culture medium containing serum.The CaCO₃(CdTe) particles could be internalized into live ceils,human liver cancer cells(HepG2) cells,for example,and most of which distributed in the lysosomes as revealed by confocal microscopy.Also the CaCO₃(CdTe) particles had low cytotoxicity in comparison with the parent CdTe QDs.Next,fluorescent silica nanoparticles with four different diameters were prepared by the classical St(o|¨)ber sol-gel method.When these silica particles were suspended into the DMEM(Dulbecco's Modified Eagles Medium)/10%FBS(fetal bovine serum) solution,some extent aggregation appeared especially for the smaller ones.Their surfaces showed a similar zeta potential of about -5mV.Cellular uptake of the silica nanoparticles with different diameters was carefully studied.The results revealed that the cellular uptake of the silica nanoparticles was dependent on particle concentration, co-culture time and cell types.While in the same condition,the cells had larger uptake amount of S370(the diameter of the particles under TEM is 368.6nm) under a 40μg/ml particle concentration.The exocytosis which is the opposite process of endocytosis was also size dependant.The silica nanoparticles could be gradually cleaned out from the cells during a 12h incubation after a 12h endocytosis,finding a higher exocytosed efficiency for smaller particles.The particles in the cells mainly distributed in the cytoplasm,lysosomes and onthe cell membranes as revealed by CLSM(confocal laser scanning microscopy).The morphology of the cells after their co-culture with particles was observed by SEM showing an aggregation state on the surface of the cell membranes for smaller ones.The silica nanoparticles were internalized into the cells through a clathrin-mediated endocytosis pathway which was proved by an obvious decrease of the cellular uptake after a low temperature incubation,NaN₃,sucrose or amandatine-HCl treatments.While the genestein treatment could not influence the cellular uptake,and the result indicated a caveolae independent endocytosis for the silica nanoparticles.The impact of silica nanoparticles on the HepG2 cells was assayed in terms of cytotoxicity and cell cycle profile.The cytotoxicity was in a low extent even at a high particle concentration up to 1000μg/10,000 cells,especially for the smaller particles.The cell cycle analysis showed that the nanoparticle-treatment can not influence the cell proliferation at a concentration of 80μg/ml for 24h co-culture.At last we synthesized porous silica particles using a CaCO₃ particle template. The porous silica particles had a spherical shape with diameters of 3～5μm and a porous structure under a dry state.Their relative surface area reached to 367.3 m²/g with multiscale pores.The concentration of TEOS(tetraethyl orthosilicate) in the preparation system showed an apparent influence on the structure of the products. Under a relative high concentration of TEOS,hollow capsules were obtained.When a PDADMAC(poly(diallyl-dimethyl-ammoniumchloride)) layer was adsorbed on the pore surface of the CaCO₃ particles in prior,more solid porous silica particles were obtained and kept the shape of the template better.The porous silica colloidal particles showed a negatively charged surface in water.Thus,TRITC-dextran (tetramethylrhodamineisothiocyanate) which is positively charged in water could be deposited into the pores.The porous silica colloidal particles could adsorb active enzymes,too.While more HRP(horseradish peroxidase) was deposited with the increase of the HRP concentration,its bioactivity was decreased.Moreover,the PSS (polystyrene sulfonate) doped CaCO₃ particles could also be used as template for the growth of silica particles on their surface.The silica colloidal particles were hollow with only a thin wall and showed a similar morphology to polyelectrolyte microcapsules under a dry state.