Preparation Of Cell Sensor Chip And Its Application In Hydrogen Peroxide Detectionin The Cells

Cell is the basic unit for morphological structure and physiological activity in live beings.Through the analysis of cell,people can be more in-depth learning and understanding the life processes.One of the cell analysis hot points is analyzing the hydrogen peroxide in cells.Hydrogen peroxide in reactive oxygen series is relatively stable,also it is very closely related to the occurrence and development of tumors.Using electrochemical sensor chip to detect hydrogen peroxide in cells can realize on-line and real time monitoring.The chip is useful in fast and sensitive for hydrogen peroxide detection.Also it can offer a new method for screening antioxidant or anticancer drugs.The main research work and results are as follows:?1?We prepare a sensor chip modified by ferriporphyrin@nano-Au which can detect H2O2.Graphene materials,gold nanoparticles and iron porphyrin materials are selected as electrode modifiers in sensor chips.Through H2O2 detection we find graphene is not useful in improving the sensor chip's sensitivity.So we choose gold nanoparticles and iron porphyrin as composite materials.Raman spectra shows that iron porphyrin materials is successfully modified on the surface of the sensor chip.We use DPV method successfully detecting the H₂O₂ which is 1×10^-6—3×10^-5 mol/L and R2 was 0.982.We also use i-t method detecting the H₂O₂ which is 1×10^-7—1×10^-6 mol/L.?2?We develop an array electrochemical sensor chip integrated with on-line culture,monitor and cell analysis.After modifying the interface,the sensor chip have good biocompatibility.And we can successfully culture the Hep3B cell.Hep3B cells were stimulated with phorbol ester?PMA?.It is found that after Hep3B cells stimulated by PMA,there is a significant increase in hydrogen peroxide in the first 4 hours.At 4 hours,the concentration of hydrogen peroxide is in the peak and it is about 0.2?M.In 4 to 6hours,the concentration of hydrogen peroxide is decreasing and the concentration of hydrogen peroxide is changing from 0.2?M to 0.13?M.It proves that the sensor can detect hydrogen peroxide releasing by Hep3B cells and quantify the amount of hydrogen peroxide production,also reveal the relationships between drug induction time and the concentration change of hydrogen peroxide.We use MTT and fluorescence detection method to verify the results.They prove that the sensor chip can achieve to detect the hydrogen peroxide in Hep3B cells.The electrochemical sensor chip can detect quick and is very convenient in cell detection.So we can conclude that the sensor chip is valuable in in-situ detection the hydrogen peroxide in cells.?3?Using electrode array cell sensor chip to culture MCF-7 and we detecte the hydrogen peroxide producing by the MCF-7 cells.We use PMA as stimulation drugs,and we find the followings.We use DPV method as our detection method and monitor the concentration of hydrogen peroxide which is coming from the MCF-7 cells for 4 hours stimulating.From the reduction peak current of DPV,we find that hydrogen peroxide increase from-34.5?A to-35?A after drug stimulation1 hour.And the concentration of hydrogen peroxide is 0.31?M to 0.36?M.After drug stimulation 2 hours,there is 0.25?M hydrogen peroxide releasing from MCF-7 cells and the reduction current is-32.5?A.After drug stimulation 3 hours,the hydrogen peroxide producing by drug-stimulated cells has basically disappeared,and the new oxidation equilibrium has been basically restored in MCF-7 cells.After drugs stimulation 4 hours,the hydrogen peroxide in cells is hardly to detect.So we can conclude that the PMA is treated in 4 hours.The experiments reveals the relationship between the induction time of drugs and the concentration of hydrogen peroxide.The hydrogen peroxide can be measured and analyzed by using hydrogen peroxide reagent kit and fluorescence labeling method.It is proved that the data obtained by electrochemical sensor chip is consistent with the results measured by electrochemical sensor chip.So it confirmes the electrochemical sensor chip can use to detection the hydrogen peroxide in reactive oxygen species from cells.