| Signal amplification technology is an important method for the detection of trace biomolecules,which thus play a significance role in the fields of biomedicine,food safety,environmental monitoring,molecular diagnostics,analytical chemistry and forensic research.Signal amplification technology is roughly divided into two categories:enzyme-assisted signal amplification methods and enzyme-free schemes.Enzyme-assisted signal amplification technology depends on the utilization of enzymes,such as polymerases,ligases,nucleases etc.These enzymes can not only specifically recognize the target sequences,but also have a high catalytic ability,but there are some shortcomings in terms of strict reaction requirements,complicated operation processes,and high costs.Therefore,the development of enzyme-free signal amplification technology approaches with mild reaction conditions and simple operation is highly demanded.In this thesis,enzyme-free signal amplification technology combined with optical biosensors for the detection of carcinoembryonic antigen(CEA),lead ions(Pb2+)were developed,and their sensing performance was systematically evaluated in terms of detection feasibility,sensitivity,selectivity,and the ability in real samples.The details are as follows:Chapter 1:some useful background information was introduced,including the clinical significance of tumor biomarkers,the current methods available for CEA detection,as well as the concept,characteristics and common application of deoxyribozyme,Other than this,several types of common signal amplification techniques were also described.In addition,the research contents and novelty of this thesis were briefly summarized.Chapter 2:A colorimetric aptasensor was constructed based on the coupling use of hybridization chain reaction(HCR)and hemin/G-quadruplex for the detection of CEA.In the sensing system,the CEA aptamer and the HCR blocker hybridized to each other and formed a double-stranded structure,which inhibits the initialization of HCR.In the presence of CEA,the aptamer specifically bound to CEA to forced the blocker strand to be released from the duplex structure,which was then ready to launch HCR.As a result,many hemin/G-quadruplex units would be generated after HCR,which upon the addition of hemin,catalyzed the oxidation of 2,2’-azino-bis-3ethylbenzothiazoline-6-sulfonic acid upon the addition of hydrogen peroxide,Meanwhile,the color of the solution changed from colorless to green,and had a strong absorbance at a wavelength of 405nm.There was a good linear relationship between absorbance and CEA concentration in the range of 30-500 ng/mL with a detection limit theorectically determined as 24.8 ng/mL.In addition,the sensor exhibited good selectivity and the recovery rates in the real serum samples was within the range of 92.2%～108.6%satisfactorily.Chapter 3:A signal amplified fluorescence sensor was developed for the sensitive detection of lead ions based on the employment of DNAzyme.In the sensing method,the hairpin strand HP-Sub was the substrate chain,and the ring region was modified with a fluorophore and quencher and the fluorescence was efficiently quenched because of the close proximity.DNAzyme was the capture chain to recognize the substrate chain by following the complementary base pairing rule.The presence of Pb2+activated the catalytic activity of DNAzymes and triggered the cleavage of the substrate chains.The fluorescence was restored due to the separation between the fluorophore and the quencher,At the same time,DNAzyme further participated in the next reaction round:cleavage of the substrate molecule followed by the signal recovery.The fluorescence was linear with the concentration of Pb2+(5 to 100 nM)with a detection limit of 3.5 nM.Moreover,the fluorescence sensor was highly specific when challenged with other divalent metal ions.The detection recovery rate of Pb2+ in tap water was 96.5%～108.9%,which provides a powerful tool for environmental monitoring and biomedical research. |
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