Research On Nucleic Acid Sensor Based On Resonant Cantilever Beam

With the rapid development of biosensor technology,a large number of sensor detection technologies have been used to specifically identify the nucleic acid sequence of the analyte to be immobilized on the interface as a capture probe,using fluorescence,electrochemistry,plasma surface resonance(SPR),Quartz microcrystalline balance(QCM),micro-electro-mechanical systems(MEMS)and other technologies to achieve the detection of the object to be tested,in the field of health and epidemic prevention,medical diagnosis,drug development,environmental science and bio-engineering and other fields have great application potential.In the development and detection of nucleic acid sensors,the composition or activity regulation of almost all nucleic acid-test substance complexes depends on the renaturation or denaturation process of nucleic acids,which is the most critical factor of interface sensing technology based on nucleic acid affinity.So far,the renaturation and denaturation kinetics of nucleic acids at the solid-liquid interface are still lack of systematic research.Therefore,the development of a new technology that is label-free,renewable,and capable of continuous online recording is very necessary for the study of nucleic acid hybridization kinetics.After more than half a century of development,MEMS sensor technology has gradually extended from the field of inertial sensors to the fields of biology and chemistry,thus providing us with the possibility of online detection of biochemical substances.We designed and manufactured an electrothermally driven piezoresistive resonant liquid phase microcantilever sensor,which is designed to reduce resonance damping and device waterproofing in the liquid phase environment.By continuously optimizing the device fabrication process and improving the detection performance,it has the characteristics of high sensitivity,stable performance,no need for labeling,fast response and real-time online data collection.It satisfies the requirements for biochemical detection of nucleic acid reactions such as the study of nucleic acid dynamic behavior at the interface.At the same time,we used SU-8 lithography to design an asymmetric curved 3D microstructure,and innovatively used the PDMS negative pressure layer to transfer the SU-8 photoresist completely to the PDMS microchannel to form a complete pattern.It is simpler to operate than traditional lithography methods and does not require expensive equipment and special materials.Then,the arcing conditions of SU-8 photoresist are explored,and the microfluidic chip with asymmetric curved microstructure is finally fabricated.Based on the microfluidic technology,we can use the PDMS microchannel to culture and separate small particles such as cells and exosomes,and it is easier and cheaper to operate than traditional separation techniques.It also lays a foundation for subsequent biological fluid sample separation and culture experiments,and further expands the detection application range of cantilever beam biochemical sensors.After completing the preparation of the cantilever beam,we made a multipolymer based interface modification system for the biochemical sensitive pool.The final optimization results in a complete modification system based on N-PDMS and polydopamine microspheres,which enables the biochemical sensitive cell of the device to fix different types of biochemical reaction materials,further expanding the detection range of the sensor.At the same time,we can detect the kinetic reaction of nucleic acid aptamer binding and dissociation with ATP through cantilever beam,and convert the frequency response signal collected by the sensor into the corresponding curve of reactant concentration and time,and then fit the nucleic acid aptamer to ATP.Affinity parameters.By comparing the published literature on the kinetic parameters of ATP-aptamers,the results of our tests are within the data parameters of the literature.The feasibility and reliability of the kinetics of nucleic acid reaction by liquid microcantilever were demonstrated.