Study On Novel Electrochemical Biosensing Technology Based On Nanomaterials

Sensitive detection of the biological macromolecules has importantsignificance in the early diagnosis and treatment of disease.Electrochemical biosensors have been widely used in detecting biologicalmacromolecule because of their unique and outstanding properties,including their high sensitive, rapid response, easy miniaturization. Butthe conventional electrochemical biosensors still have numerousproblems such as low sensitivity and poor stability, which is difficult tomeet the requirements of clinical medicine. In this dissertation, combinedwith nano-materials and nucleases, put forward three types ofelectrochemical biological sensing technology in detecting biologicalmacromolecules. The main research are listed as follow:(1) A novel label-free gap-electrical biosensor strategy based onself-catalytic growth of unmodified AuNPs as conductive bridgesmediated electrical signal transduction for DNA hybridization detectionhas been developed. In this strategy, the presence of target DNA canmediate the variation of the GOx-like catalytic activity of AuNPsassembled onto the insulative gaps, attributing to the differentelectrostatic interactions between AuNPs and single-stranded (ssDNA)and double-stranded (dsDNA), respectively. This electrochemicalbiosensor has been demostrated to exhibit sensitively response to thetarget DNA concentation range from100pM to1μM with the detectionlimit of100pM.(2) A new homogeneous electrochemical biosensing technology forT4PNK detection has been proposed, based on the conductivity ofgraphene and the Auxiliary signal amplification of exonucleaseⅢ. In this method, PNK transfers the terminal phosphate of ATP to5’-hydroxyltermini of DNA, then OligoA and OligoB could be ligated by DNA ligseand formed a stable duplex with ferrocene labelled HP probe. In thepresense of exonucleaseⅢ, it digests the probe into mononucleotidsincluding a ferrocene labelled electroactive mononucleotide, the lowaffinity of the mononucleotide to graphene resulted in no electrochemicalsignal. In the absence of T4PNK, the ferrocene labelled HP probe couldbe attracted by graphene resulted in high electrochemical signal. Thestrategy has been demostrated to exhibit dynamic responses to the T4PNKconcentration range from0.0001UmL-1to1UmL-1with the detection limitof0.0001UmL-1.(3) Taking the advantages of the strategy in the chapter3, anothernovel homogeneous electrochemical biosensing technology based on theconductivity of graphene and the Auxiliary signal amplification ofexonucleaseⅢ for analysising dam has been exploited. In this method,the dsDNAs containing the methylated site-specific were methylated bythe dam and dissected to fragments by Dpn Ⅰ, triggering thehybridization with the ferrocene labelled P3and P4. In the presence ofexonuclease III, the labelled ferrocene electroactive probes have beendigeted into mononucleotides, resulting in the low affinity to thegraphene and no electrochemical signal generated. The strategy has beendemostrated to exhibit dynamic responses to the dam concentration rangefrom0.01UmL-1to30UmL-1with the detection limit of0.01UmL-1.