Research On Microfluidic Chip For Single-cell Analysis

After years of development,microfluidic technology has been used in chemistry,physics,biology,medicine,materials science,optics and Micro-electromechanical Systems,and has become an important interdisciplinary topic.Some typical applications,such as real-time detection,cell sorting,and cell analysis,bring benefits to life and researches.Microfluidic technology provides efficient and rapid methods to analyze biological cells in life science.The dimensions of microfluidic devices are in the same scale with the size of cells and microbes.Thus,microfluidic systems enable the manipulation and analysis of a small amount of cell populations or even single cells.In this thesis,a background of microfluidic technology and a summary of previous methods of single-cell trapping and culturing has been stated in the first chapter.In order to address the existed questions of current methods,we have designed a microfluidic array for single-cell trapping based on Hagen Poiseuille's law.A flow resistance model of channel network has been developed in MATLAB for design optimization.Based on MATLAB theoretical model,a microfluidic chip with 96 cell traps has been designed and fabricated.These 96 traps have been patterned in an array format(16-row x 6-column).Each trap has been constructed as a narrow slit between two fluidic channels,namely a main-flow channel and a side-flow channel.Cells can be captured and retained at the ends of these narrow slits based on the hydraulic pressure drop across the slits.The pressure drop across the cell traps in each column can be controlled by adjusting the flow rates of the main-flow and side-flow channels,so that adequate pressure drop for cell immobilization can be generated.In the microfluidic chip,6 parallel columns have been designed for the comparative cell culturing,and each column has been patterned with 16 cell traps.In order to deliver samples to the 6 parallel columns in sequence,3 inlets have been designed to perform the laminar-flow perfusion.By changing the flow rate ratio of the inflows from the 3 inlets,cell samples can be guided to the 6 columns of cell traps in turn.After cell immobilization in all 6 columns,comparative cell culturing has been conducted by exposing the immobilized cells in various columns with different cell-culturing media.The processes for chip fabrication are as follows:First,channel masters have been fabricated on SU-8 photoresists through photolithography.Then,PDMS fluidic channels have been replicated from the SU-8 masters by using soft lithography process.After the surface modification of two PDMS layers through O2 plasma we can assemble them irreversibly and obtain a double layer of PDMS channel network.Finally,the PDMS channel network has been sealed with a glass slide.Then three experiments have been designed and carried out to verify the functions of this microfluidic system:(?)Laminar-flow perfusion has been verified by using dye solution delivered to 6 parallel channels in sequence;(?)Beads have been used to characterize the trapping efficiency of this microfluidic system.(?)Yeast cells have been used for single-cell immobilization and comparative culturing on this microfluidic system.The results of experiments have shown that our microfluidic single-cell culturing system enables single-cell immobilization in turn by laminar sheath-flow focusing,a high efficiency(85%)of single-cell capture,and comparative cell-culturing on a single chip.