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Real time monitoring single cells by microelectrodes array
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本研究使用阻抗分析觀察量測區域內的電特性，並成功建立出符合其物理意義的等效電路模型，以實際量化其阻抗量測結果。首先，將不同的溶液(PBS以及培養液)注入晶片中觀察其阻抗特性，之後將三種不同種類的細胞(肺腺癌細胞A549、動物腎上皮細胞MDCK以及子宮頸癌細胞HeLa )分別注入晶片中，並使用阻抗分析儀來量測細胞阻抗；其頻率範圍選用600 Hz~100 kHz、電壓輸入範圍選用0.1 V~1.0 V 進行定電壓阻抗量測，藉由其阻抗量測結果可以明確分辨出不同種類的細胞。利用其等校電路模型之擬合運算，可以詳細得知細胞相對應得電子元件數值，如細胞膜電阻、細胞膜電容與細胞質電阻。當量測電壓為1.0 V時，相較於0.7 V之阻抗量測結果會有明顯降低的情形，且等效電路擬合之相位角出現了不吻合的情況。因此我們推論，輸入的電壓過大將會導致細胞膜表面有些微的穿孔現象發生，故本研究最後使用微流電極晶片產生細胞電穿孔效應，並同時使用阻抗量測與細胞染色來觀察驗證細胞膜穿孔前與穿孔後之差異。
In this thesis, we present a microfluidic device with microelectrodes array which is capable of measuring the impedance of single cells and providing electrical impulse for electroporation of single cell. Single cells are captured in microstructures using fluidic transmission. The device includes a glass substrate with electrodes array and PDMS with main channels, branch channels and capturing areas. An equivalent circuit model of flow resistance is used to study the flow of microfluidic device. According to analytical results, single cells are able to be captured in specific areas experimentally.
In this study, the impedance of single cells and medium is successfully measured by impedance spectroscopy. An equivalent circuit model of impedance is established and fits closely to the experimental results. The system is operated at frequency between 600 Hz and 100 kHz, and the operating voltage is from 0.1 V to 1.0 V. The resistance and capacitance of cell membrane and the resistance of cytoplasm are measured by the simulation of equivalent circuit model of impedance.
Finally, the electrodes array is used to provide the electrical pulse to cell for electroporation. The amplitude of electrical impulse 2.2 Vpp may be used to open the ionic channels of the cell membrane for the future application of electroporation. The cell electroporation is observed both by optical microscope to record the entrance of Trypan blue stain and impedance measurement.
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