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Title: Design and fabrication of high-throughput microfluidic device for cell-based assays
Other Titles: She ji he zhi bei yong yu xi bao shi yan de gao tong liang wei liu jing pian zhuang zhi
Authors: Li, Cheuk Wing (李卓榮)
Department: Dept. of Biology and Chemistry
Degree: Doctor of Philosophy
Issue Date: 2006
Publisher: City University of Hong Kong
Subjects: Biological assay
Fluidic devices
Notes: CityU Call Number: QH581.2.L526 2006
Includes bibliographical references.
Thesis (Ph.D.)--City University of Hong Kong, 2006
xiv, 190 leaves : ill. (some col.) ; 30 cm.
Type: Thesis
Abstract: Microfluidic systems have shown unique advantages in performing analytical functions such as controlled transportation, immobilization, and manipulation of biological molecules and cells, as well as separation, mixing, and dilution of chemical reagents, which enables the analysis of intracellular parameters and detection of cell metabolites, even on a single-cell level. However, traditional microdevice fabrication based on silicon technology requires expensive and specialized equipments. In order to reduce the fabrication cost, we have developed two methods to prepare microfluidic devices with multi-height structures using single step photolithography. Specifically, two types of commercially available transparencies were employed as photomasks to generate master based on conventional printed circuit board (PCB) technology. Multi-height microstructures can be produced by different patterns featured on transparencies. For example, in thermal color-printed transparency, patterns of different light opacity were used to generate multi-height microscale structures while “windows” and “rims” patterns were printed on film transparencies to produce microtunnels after isotropic wet etching. Poly(dimethylsiloxane) (PDMS) was then moulded against the positive relief PCB master to generate microfluidic structures. These simple microfabrication processes require only low-cost materials and minimal specialized equipment and can reproducibly produce microchannels with dimensions that are sufficient for most microfluidic applications. Moreover, these microfabrication methods offer high throughput production (multiple devices on 8 in.12 in. PCB board) capability that surpasses the throughput based on traditional silicon technology (3 in. silicon wafer). We have demonstrated the quality of the microfluidic devices by performing basic laminar flow characteristics in microchannels, controlled immobilization of biological cells on the multi-height structures together with on-chip immunocytochemical staining assays. The mentioned microfabrication methods enabled flexible and cost-effective fabrication of microfluidic structures, including but not limited to trap biological cells. In order to immobilize biological cells with high efficiency and easy control while maintaining cell viability, one approach for cell immobilization is to utilize constriction structures such as dams to trap cells in microfluidics. So we further our studies by developing computational models to analyze two different types of constriction structures for cell immobilization: dams either in perpendicular or in parallel to the main flow route. Various structural models and experimental conditions were compared for cell docking and alignment, and the pressure and velocity profiles of the flow in the micro-channels and the hydrodynamic force and shear stress on the docked cells were calculated based on fluid dynamic theory and numerical simulation. The effects of the dam structures and cell docking on the flow properties, the transportation efficiency, and the induced stress on the docked cells were analyzed. Improved hydraulic pressure profiles in the auxiliary inlets were discussed for the modulation of the flow characteristics and attenuation of hydrodynamic forces exerted on the cells. Together with microfluidic technologies in controlling cell transportation, generating concentration gradients, and monitoring cellular responses, our low-cost microfabrication method and improved cell docking provide a strong foundation to establish an integrated, high-throughput microdevice for bioassays. We have developed microfluidic modules featured with channel components and sandbag structures for positioning biological cells within the microchip. We have demonstrated that by geometric modulation of the microchannel architectures, it is possible to immobilize individual cells at desired locations with controllable numbers, to generate defined concentration gradients at various channel lengths, and to improve the efficiency and reproducibility in data acquisition. The microfluidic module was used to exercise a series of cell-based assays, including the measurement of kinetics and dynamics of intracellular enzymatic activities, the analysis of cellular response under the stimulation of two chemicals with defined concentration profiles, and the study of laser irradiation effect on cellular uptake of photo-sensitizers. The results demonstrated the capabilities of the microfluidic module for conducting multiple sets of dose-dependent, cell-based bioassays simultaneously and for comparing responses of individual cells under various stimulations quantitatively.
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