Construction Of Porous Nanocomposites And Its Application In Biomolecule Detection

Since tumor biomarkers present in body fluids or tissues are usually expressed at low levels in the early stages of disease,traditional clinical assays are difficult to achieve highly sensitive detection of these tumor biomarkers.Therefore,it is of great importance to develop and design novel ultrasensitive detection strategies and bio-probes.The emergence of porous nanomaterials provides a new class of application tools for the detection and imaging of tumor markers.In this thesis,a series of fluorescent detection platforms were constructed using porous nanomaterials by combining the advantages of fluorescence analysis and detection,and endowed with additional functions and properties through the modification and functionalization of various biomolecules.Meanwhile,in order to improve the sensitivity of the constructed probes,this paper combines signal amplification techniques such as controlled release,enzyme-assisted target cycle amplification strategy and aggregation-induced enhancement effect to achieve highly sensitive detection of low-expression tumor biomarkers as well as cell imaging.The main studies includes:(1)A dual amplification controlled-release fluorescent probe based on the combination of gold nanocage(Au NC)and nucleic acid exonuclease III(EXO III)and used for cellular telomerase activity detection was designed and developed based on controlled-release and enzyme-assisted target cycle amplification techniques.First,single-stranded nucleic acid molecules capable of triggering the telomerase amplification reaction are assembled as gating units on the Au NC surface by electrostatic interaction to achieve controlled release of the dye molecules.Compared with other conventional fluorescence detection strategies,the Au NC-based fluorescent probe can achieve a selective response to telomerase with a simple modification.More importantly,the probe does not require any external stimuli(e.g.,radiation or p H),and only requires telomerase to trigger the gating switch to release a large number of signal molecules,which is enhanced and accelerated by the enzyme-assisted target cycle amplification technique,thus achieving a highly sensitive detection of telomerase activity.Thus,this controlled-release fluorescent nanoprobe provides a novel strategy for the detection and in situ imaging of telomerase activity.(2)A multifunctional polydopamine(PDA)-coated metal-organic backbones(MOFs)biosensor based on Zn²⁺-triggered aggregation-induced enhancement effect(AIE)is proposed for the highly sensitive detection of mi RNA-122 and synergistic chemo-photothermal therapy in vitro.Firstly,a layer of PDA was modified on the surface of ZIF-8 particles loaded with doxorubicin hydrochloride(DOX)by a self-polymerization reaction.on the one hand,the introduction of PDA increased the biocompatibility of the nanocomposite carriers;on the other hand,the coordination between the catechol group of PDA and the metal ions of ZIF-8 accelerated the decomposition of the ZIF-8 structure in an acidic environment and promoted the Zn²⁺and DOX release efficiency.Meanwhile,the AIE signal amplification strategy is driven by the metal cofactors released from the collapsed MOFs structure,avoiding cellular damage caused by enzyme inactivation and enzyme transport.In addition,the MOFs structure has high drug-carrying capacity and the PDA has photothermal conversion properties,giving the biosensor additional functions,including synergistic chemo-photothermal therapy.Thus,self-sufficient multifunctional PDA-coated MOFs biosensors provide a scientifically intelligent and ideal strategy for the development of integrated nanocomposite carriers for early cancer diagnosis and treatment.(3)A fluorescent probe based on gold nanoclusters(Au NCs)and metal organic backbone(MOFs)nanomaterials was designed for the ratiometric detection of mi RNA-21.Ui O-66-NH₂ can be used to load more Au NCs and probe molecules due to its high specific surface area,high porosity,and the abundance of easily functionalized amino groups on the surface.Second,the construction of ratiometric fluorescent probes was achieved by covalently cross-linking the designed probe molecules onto the surface of Au NCs@Ui O-66-NH₂.When the probe entered the cell through cytosol,the overexpressed mi RNA-21complementarily hybridized with the single-stranded DNA of the modified FAM and detached from the surface of Au NCs@Ui O-66-NH₂,resulting in the recovery of the fluorescence signal of the FAM burst by BHQ,while the fluorescence signal of Au NCs in Au NCs@Ui O-66-NH₂ remained unchanged.Thus,ratiometric detection of mi RNA-21 was achieved by the ratio of the recovered fluorescence signal of FAM dye to the fluorescence signal of Au NCs.Meanwhile,MCF-7 breast cancer cells and L-O2 normal hepatocytes were used as a model to confirm that the probe can be used for the differentiation of different mi RNA-21 expressing cells,which provides a potential application tool for clinical diagnosis.