| Exosomes are the smallest subset(30-150 nm)of extracellular vesicles(EVs)produced by all endocytic cells and stably exist in all types of body fluids.They carry a variety of biological macromolecules from their parental cells,including proteins,lipids and nucleic acid molecules,and serve as an intercellular communication system for cell long-distance communication.Due to the important role in promoting tumor growth and metastasis,exosomes have great research values in the field of tumor diagnosis and therapy.Especially,tumor-derived exosome subpopulations expressing tumor-specific markers are considered to be biomarkers for tumor liquid biopsy with broad application prospects in the field of clinical diagnosis.Biosensing technology is expected to achieve rapid point-of-care testing of exosomes due to its high efficiency,sensitivity,specificity with smart and economic device.The fluorescent nanosensor based on nano-materials can realize low-abundance molecular liquid in complex biological samples owning to the characteristics of strong anti-interference,less sample consumption and simple operation,which makes it possible to rapidly detect low-abundance tumor-derived exosomes subpopulations.Aggregation-Induced Emission(AIE)probes highly dispersed in solution have extremely weak fluorescence;while strong fluorescence is emitted in the aggregate state.Due to its good solubility and biocompatibility,AIE probes are suitable for the construction of "Turn-on" liquid-phase detection methods based on fluorescence nanobio sensing technology.In the first chapter of this thesis,we identified the PSMA(+)exosome subpopulation of prostate cancer by PSMA aptamer,and successfully constructed the PSMA(+)exosome subpopulation fluorescence nanobiosensor detection technology for prostate cancer based on "Turn-on" fluorescence reporting system consisting of the fluorescent reporter group AIE probe TTAPE and the fluorescent quenching group graphene oxide(GO).It can realize the quantitative detection of PSMA(+)exosome subpopulation of prostate cancer.On this basis,we optimized the reaction conditions,and evaluated the detection performance of the method,such as specificity,sensitivity,and linear range.Finally,the PSMA(+)exosome subpopulation of prostate cancer in clinical plasma samples was tested to evaluate the clinical application value of this method.In the second chapter of this thesis,we used GPC-1 antibody functionalized magnetic beads as microcarriers for GPC-1(+)exosome subpopulation isolation and biosensing detection.The "sandwich" type recognition system composed of GPC-1 antibody on the surface of magnetic microcarriers and CD63 aptamer was adapted to specifically identify GPC-1(+)exosome subpopulation of breast cancer.And the recognition signal was transformed into the formation of DNA nanostructure by the link between ssDNA and the CD63 aptamer.To increase the sensitivity of this method,a "Y" shape DNA self-assembly nanostructure amplification system and TTAPE/GO fluorescence reporting system mentioned above were used to construct exosome fluorescence nanobiosensor detection technology to realize the ultracentrifugation-free quantitative analysis of GPC-1(+)exosome subpopulation of breast cancer.On this basis,we optimized the reaction conditions,and evaluated the detection performance of the method,such as specificity,sensitivity,and linear range.Finally,the GPC-1(+)exosome subpopulation of breast cancer in clinical plasma samples was tested to evaluate the clinical application value of this method.In summary,two kinds of fluorescence nanosensing systems were successfully constructed and applied to the liquid-phase detection of prostate cancer and breast cancer derived exosomes respectively,which provided experimental and theoretical basis for the construction of new methods for rapid and simple cancer liquid biopsy. |
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