Detection Of Biomarkers Based On Graphene Oxide And Fluorescent Silicon Nanomaterials

With the rapid development of the society and economy,people have shifted their attention to human health and life sciences.The continuous development of life sciences requires and urges researchers to study the life process at the molecular level and carry out some early diagnosis and treatment to related diseases,which undoubtedly poses an unprecedented challenge to analytical chemistry,especially for biochemical analysis.Obviously,the traditional analytical methods and technical means can not meet the needs of further research in the field of life science.As a new biological analysis method-biosensors,in virture of its good selectivity,high sensitivity,low cost,simple operation and fast response and so on,have been developed rapidly in the recent years in the field of analysis science.At present,biosensors have been widely used in the analysis and detection of biological macromolecules(nucleic acids,proteins,enzymes and peptides,etc.),biological small molecules,heavy metal ions,biological drugs and so on.Although biosensors still have many shortcomings in improving the ability of anti-interference in practical applications and real-time monitoring,we believe that with the development of science and technology,especially nanotechnology,materials science and the rise of biochemical analysis of interdisciplinary,biosensors will inevitably be widely used in drug development,environmental monitoring,health care and so on.Based on the new DNA-crosslinked graphene oxide and two-photon fluorescent mesoporous silica nano-probes,we developed two new type of nano-sensing methods for combinatorial cancer therapy and detection of GSH activity.The main contents of the thesis are summarized as follows:Graphene oxide(GO),a shining star in the field of materials science,has many advantages in combinatorial cancer therapy:non-toxic,easy to function by covalent and non-covalent methods,photothermal effects,and high drug/biomolecular loading efficiency also enables it to be used in real-time chemistry and photothermal processing.Due to these advantages,many methods based on graphene oxide have been developed for combining cancer treatments.The pH response has unique advantages due to the difference in pH between normal and tumor tissues,so pH can be an excellent method of cancer targeting.A proton-powered DNA nanostructure made up of four Cytosine-rich C's is a good choice for cancer targeting.This DNA sequence forms a closed four-chain i-motif structure under weak acid conditions and folds arbitrarily under neutral to alkaline conditions.This conformational change can be accomplished within seconds to tens of seconds so that low pH in tumor cells and tissues is considered to be an ideal trigger for the selective release of anti-cancer drugs in tumor tissues and cells.In Chapter 2,we first assemble a ssDNA probe,which is rich in guanine(G),and aptamer sgc8,which targets protein tyrosine kinase 7(PTK7),by covalent cross-linking into PEGylated Graphene oxide on the surface.The fluorophore-labeled probe sequence was then synthesized using the 5'-end i-motif sequence and the 3'-end DNA methyltransferase(DNMT1)capture sequence.In a neutral pH environment,a fluorophore-labeled probe can be aggregated in poly(poly)-polyglucoside by hybridization with a graphene oxide-rich,guanine-rich single-stranded DNA probe Glycol-modified graphene oxide on the surface.After hybridization,the fluorescence of the fluorescent probe is highly quenched due to the high quenching efficiency of graphene on the fluorophores thereon.In addition,as a model anti-cancer drug(DOX),it can easily be loaded into hybridization complexes by simple mixing.The surface of nanocomposites is modified with polyethylene glycol to encourage nanocomplexes to be efficiently endocytosed by tumor cells.The low pH in tumor cells triggers the formation of an i-motif structure,accompanied by dissociation of the probe from the graphene oxide surface.Disassociation of the probe results in the release of DOX,activating a strong fluorescence response and inhibiting DNA methyltransferase activity by covalent capture.Our polyethylene glycol-modified graphene oxide DNA nanocomposites(GO-PEG-DNA)contain the functions required for cancer treatment and on-demand drug delivery compared to currently available systems.Glutathione(GSH)provides important cellular biology functions and its abnormal levels are associated with many diseases.Therefore,the establishment of high selectivity,high sensitivity simple and rapid method for the determination of glutathione is of great significance.The imaging of optical organisms stained with fluorescent probes may be the most attractive technique for detecting bioreductive species in vivo due to their high sensitivity,real-time spatial imaging and the non-destructive detection of targets in biological systems.In Chapter 3,we developed a newly prepared fluorescent silicon nanoparticle probe for monitoring GSH as a fluorescence reporter and manganese dioxide(MnO2)nanoplates as fluorescence quenchers and recognition units.The newly prepared fluorescent silicon nanoparticle probes have a positive charge due to the presence of amino groups on their surface and can easily adsorb negatively charged MnO2 nanosheet by electrostatic interaction.Due to the strong quenching ability of MnO2 nanosheets,the fluorescence of the fluorescent silicon nanoparticle probe is easily quenched by MnO2 nanosheets.However,the introduction of GSH selectively activates and significantly enhances the fluorescence of the fluorescent silicon nanoparticle probe because GSH can reduce MnO2 nanosheets to Mn2+ and lead to the decomposition of MnO2 nanosheets.Fluorescent silicon nano-probes showed high sensitivity to GSH,reaching a detection limit of 200 nM.It also exhibits high selectivity for GSH relative to other biomolecules and electrolytes,with good membrane permeability and excellent biocompatibility.Most importantly,it has been successfully applied to intracellular fluorescence imaging of GSH in living cells and tissues.