Nanomaterial And Nucleic Acid Probe-based Fluorescent Biosensors

DNA and proteins are the biomarkers that have critical applications in many areas of clinical diagnostics like early cancer diagnostics, infectious disease diagnostics, prenatal diagnostics, treatment monitoring and personalized medicine. The significance of biomarkers detection drives the demand for developing sensitive, specific, simple and cost-effective biosensors. A fluorescent biosensor for biomarker detection that based on the transformation of biomolecules recognition into fluorescent signal possesses the advantages of sensitive, simple and rapid and has been widely used for biomolecules detection. The rapidly developed nanotechnology brings the nanomaterials with special optical properties into fluorescent biosensors, which improves the performance and promotes the development of fluorescent biosensors. Nanomaterials based fluorescent biosensors becomes a new direction of the development of biomolecule analysis.Here two fluorescent biosensors based on tungsten disulfide(WS2) and graphene oxide(GO) nanosheets, respectively, had been developed for DNA and interferon-gamma(IFN-γ) detection. The nanosheets were used as quenchers and detection platforms due to their large surface area, different binding affinity to different structure of nucleic acids and high fluorescence quenching efficiency. Peptide nucleic acid(PNA) and aptamer with high binding affinity and specificity were employed as the probes in the biosensors. The major contents were as follows:In chapter 1, the WS2 nanosheet and PNA based fluorescent biosensor had been developed for rapid DNA detection. In the absence of DNA, the PNA probe was absorbed on the surface of WS2 nanosheet and fluorescence quenched efficiently. When PNA probe was hybridized with DNA, the PNA-DNA duplex formed and had weak interactions with WS2 nanosheet, thus the fluorescence remained. The biosensor could detect DNA sensitively and specifically by measuring the fluorescence intensity.In chapter 2, the aptamer and enzyme-aided amplification based fluorescent biosensor had been developed for IFN-γ detection by using GO nanosheet as quencher and detection platform. The hairpin probe HP1 consisted of the aptamer sequence of IFN-γ. When IFN-γ bound with the aptamer region, the hairpin structure of HP1 was opened and a single stranded DNA sequence of HP1 exposed. The exposed DNA sequence could hybridize with the DNA fragment at terminal of HP2 probe and triggered the exonuclease III(EXO III) aided amplification, resulting in the degradation of HP2 and release of single stranded DNA template. The DNA template hybridized with the fluorescent DNA probe and triggered the EXO III cleavage of probe, leading to the DNA template recycling and fluorophore release. Large amounts of fluorophores were obtained after many catalytic cycles. The introduction of GO could not quench the fluorescence of fluorophore efficiently and the fluorescence intensity was very strong as the affinity between GO and fluorophore was weak. In the absence of IFN-γ, the hairpin structure of HP1 could not be opened and the catalytic reaction would not occur. Upon the addition of GO, the fluorescent probe absorbed on the surface of GO and the fluorescence was quenched. IFN-γ could thus be detected by the biosensor via fluorescence measurement.