| A variety of therapeutic approaches including chemotherapy,molecularly targeted therapy,gene therapy,radiotherapy,immunotherapy and phototherapy have been extensively applied to treat cancers in clinic.Nanomedicine provides an innovative diagnostic or therapeutic regime for precise cancer treatment,because nanomaterials can be flexibly designed to provide a high-performance multi-functional nano-drug carrier to satisfying drug delivery.Unfortunately,the majority of the nanoparticles intratumoral were either trapped in the extracellular matrix or taken up by perivascular tumor associated macrophages.The low cancer cell targeting efficiency and significant uptake by normal cells suggest the need to re-evaluate the active targeting process and therapeutic mechanisms using quantitative methods.This will be important for developing strategies to deliver emerging therapeutics such as genome editing,nucleic acid therapy,and immunotherapy for cancer treatment using nano-drug delivery.Therefore,there is an urgent need for a simple,rapid,and universal quantitative analysis method to achieve highly sensitive quantitative analysis of nano-drug delivery.This study focus on to design based-nanomaterials fluorescent probe as a marker to quantify nano-drug delivery using high specificity,high sensitivity and high throughput fluorescent immunoassays.1.Preparation and identification of polymer nano-drug delivery haptens,immunogens and antibodiesA spherical nano-drug delivery hapten(PSIOAm)was prepared by ultrasound.In view of the carrier protein enhancing the immunogenicity of the nanoparticles,the preparation of a nanocarrier immunogen(PSIOAm-BSA)with BSA carrier protein and a coated antigen(PSIOAm-OVA)with ovalbumin was carried out.The polyclonal antibody against the nanocarriers with a titer of 1/64000was successfully obtained by immunizing New Zealand white rabbits with immunogen.2.An evaporation induced self-assembly approach to prepare polymorphic carbon dot fluorescent nanoprobes for protein labellingCarbon dots are one of the most promising fluorescent probes in the biomarker field,because of the high fluorescence quantum yield,high anti-photobleaching and non-toxicity.In this experiment,a green non-toxic carbon dot as a precursor and glutaraldehyde as a self-assembly initiator.Combined with evaporation-induced self-assembly technology,carbon nanoclusters were studied at pH=7.4,0.1M NaCl,0.01M PBS and 0.1 M PBS effect on the self-assembly morphology of carbon nano-clusters.Carbon-based nanospheres with better dispersibility formed at pH=7.4,Carbon-based nanowires were formed in 0.1 M NaCl,and carbon-based nano-belts were formed in 0.01 M PBS and 0.1 M PBS with good dispersibility were successfully synthesized.A new method for the preparation of based-carbon fluorescent probes with different morphologies has been developed to effectively control the concentration and type of inorganic salts.Under conditions of forming spherical carbon dot nanoprobes at pH=7.4,Self-assembly between carbon nanoclusters and proteins,successfully prepared novel carbon dot fluorescent probe labeling protein,and optimize the optimal labeling amount of protein.3.Carbon dot-linked fluorescent immunoassay detection polymer nano-drug deliveryThe carbon dots have higher anti-photobleaching and no fluorescence flickering characteristics than conventional fluorescent probes.In quantitative analysis experiments,carbon dot fluorescent probes can increase the sensitivity of the experiment and the reliability of the measured signal values.In this experiment,the self-assembled carbon dot as a signal collection probe,establish a carbon dot-linked fluorescent immunoassay to quantify the polymer nano-drug delivery.Under the optimal conditions,the standard curve was F=13955.08-981.93lgC.The correlation coefficient was R2=0.994,the linear range was 5×10-4-5×102 ng/mL,the detection limit was 0.15pg/mL,the standard addition recovery was85%-103.6%,cross-reaction experiments were carried out on materials which similar and different compositions with analytes,all not exceed 0.01%.Developed a new high sensitivity,high throughput and high specificity method for quantification of polymer nano-drug delivery.4.Magnetic probe-linked fluorescent immunoassay detection polymer nano-drug deliverySquare ferroferric oxide?Fe3O4?synthesized by high temperature co-precipitation,and successfully transformation into aqueous with CTAB.With bovine serum albumin as the carrier of isothiocyanate fluorescent,hydroxypropyl cyclodextrin?CD?as an anti-quenching agent and increase the electronegativity of fluorescein carrier,reserch on effects of different types and concentrations of hydroxypropyl cyclodextrin on the fluorescence intensity of fluorescein carrier,magnetic fluorescent probe obtained based on electrostatic interaction.After silica modification,obtain a based-silica magnetic fluorescent probe with good biocompatibility and high brightness.A preparation method for a high-brightness magnetic fluorescent probe has been developed,which convenient for separation and purification of fluorescently labeled proteins.Based on the above,establish silicon-based magnetic probe-linked fluorescent immunoassay to quantify the polymer nano-drug delivery.Under the optimal conditions,the standard curve was F-F0=11477.59-1537.63lgC.The correlation coefficient was R2=0.994,the linear range was 5×10-5-5×103 ng/mL,the detection limit was 3×10-7ng/mL,the standard addition recovery was 97%-112%,cross-reaction experiments were carried out on materials which similar and different compositions with analytes,all not exceed 0.01%.Compared with the carbon dot fluorescence immunoassay,this method makes full use of the magnetic separation characteristics of the based-silicon magnetic fluorescent probe and the red-emitting luminescent probe to develop a simple,efficient and highly specific quantitative polymer nano-drug delivery. |
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