Construction Of Biochip Based On Dielectrophoresis And Its Application Research In Cellular Assays

The biochip technology has attracted considerable attention since it came out due to its as fast speed, less sample consumption, high sensitivity and ease of integration and automation. As one of the important biochips, the dielectrophoresis (DEP) chip refers to the electrokinetic effect resulting from a polarizable particle under the action of a non-uniform alternating current (AC) electric field and achieves to manipulate, transport and separate different bio-particles. According to the configuration of micro-electrodes, they can be divided into three types: conventional dielectrophoresis (cDEP) chip, traveling wave dielectrophoresis (twDEP) chip, electrorotation (ROT) chip. In the past decades, chip-based dielectrophoresis technology has become a powerful analytical and detective method to detect, manipulate and fractionate cells and other bio-particles since it offers many advantages, including non-invasive detection, high sensitivity and fast speed.The work presented in this thesis consists of three parts. One is the implementation, optimization, and application of the ROT chip for cell detection; the second is the construction and application of the twDEP chip and the third is the study of the automated cell fractionation system.In the setup, two configurations of the ROT chip were developed and fabricated based on the research of the basic theory and principle of design of the DEP technology. An experimental platform was built up and a ROT array chip was developed to improve the throughput and efficiency. The distributions of the electric field and the DEP force (torque) were simulated when the micro-electrodes worked in"DEP mode"or"ROT mode"by using finite-element-analysis (FEA) method, respectively. The DEP force assisted positioning technology and the ROT technology were integrated to manipulate and detect cells for the first time and were validated by experiments.By using this setup, the electrokinetic responses of autotrophic and heterotrophic micro-algal cells with different biochemical components were researched for the first time, as well as the dielectric alterations of the apoptotic HL-60 cells induced by bufalin. The results demonstrated the dielectric properties can be monitored by our chip and the specific membrane capacitance of the HL-60 cells fell from an initial value of 15.6±0.9 mF/cm2 to 6.4±0.6 mF/cm2 after a 48 h treatment with 10 nM bufalin. However, the average membrane conductance remained almost constant during the first 24 h of bufalin treatment and then increased distinctly thereafter.According to the dielectric alterations detected by the ROT setup we have constructed, a cell separation chip was designed and developed based on a combination of the principles of twDEP and laminar flow. The traveling wave electric field and the laminar flow field were simulated. A twDEP chip was fabricated by patterning parallel Au/Cr electrodes, and bonded with a PDMS microfluidic channel after oxygen plasma treatment. By using this chip, the separation of Jurkat cells from human red blood cells in the mixture was realized. The result demonstrated a displacement of 300μm differing from each other was achieved within 1 min.To improve the efficiency and automatization, we present a idea of automated cell fractionation system. The transparent ITO conductive glass was used to fabricate the micro-electrode instead of conventional Au/Cr electrodes to form the twDEP chip, which greatly reduced the background noises generated during experiments. At the same time, a software was developed for cell motion measurements and a real time, on line monitoring system was realized during the cell separation processing. The result demonstrated more than 90% cells can be recognized by the system and the error was no more than 5%.