Research On Cells Separation Device Integrated With Micro-flow-regulator

Blood samples are important objects for biomedical detection.Blood samples usually need to be pre-treated（removal of cells or retaining certain cells）.Traditional method to obtain plasma or other components through centrifugation often has problems,such as low purity,high cost,and the need for professional operation techniques.This thesis mainly focuses on the targets in the blood realizing the pretreatment of blood samples through microfluidic technology,including plasma separation and the sorting of circulating tumor cells.We have studied the flow regulator based on microvalves,and realized a low-cost,handheld sorting device by integrating the micro-flow regulator and the inertial micro-chip.The research results are as follows:（1）A low-cost,rapid prototyping beehive-like multilayer polymer film chip device was developed,which could be used for ultra-high-throughput plasma separation.In the first step,the influence of the flow channel cross-sectional size and the input flow rate on the particle focusing behavior was studied.Then,blood samples with different dilution ratios were injected into the selected flow channel to verify the feasibility of its application in plasma separation.The plasma separation efficiencies of diluted blood samples with HCT of 0.5%and1%were 98.9±0.25%and 96.7±1.0%（n=3）,respectively.Finally,the beehive-like multilayer polymer chip was designed and manufactured to realize ultra-high-throughput plasma separation.For a 50-fold diluted whole blood sample,the total processing volume can reach up to 72 m L min^-1,and the separation efficiency is about 90%.（2）The flow regulation characteristics of microvalves with various structures were studied;the influence of different parameters on the flow regulation was studied.An eight-channel parallel shut-off valve structure was designed.After testing,the microvalve could maintain linear change of flow rate in the low-pressure stage;after reaching the threshold pressure,it quickly dropped to zero,and the microvalve shut down.A low-throughput flow regulator valve was studied.The structure design was optimized to fabricate an all-polymer-film chip which improves the repeatability of the microvalve constant flow rate.the double membrane layers flow regulator was studied.The influence of the width of the bypass channel on the flow regulation characteristics was studied;based on the study of double membrane layers flow regulator,the fabricating process and structure design were simplified.A single membrane layer deformed flow regulator was proposed.The effect of PDMS film,main flow channel height,fluctuating pressure input and other parameters on the flow characteristics was studied;the flow channel structure was further optimized to solve the problems of bubble and repeatability;in addition,the multi-level arrangements of flow regulator were designed and tested.In the end,the problem of single-membrane-layer deformation was successfully solved through the shape change of the deformation area,so that the flow rate regulator could meet needs for the subsequent integration.（3）A handheld instrument for sorting rare circulating tumor cells was designed and manufactured.First,the feasibility of integrating the single-inlet spiral chip for plasma separation and the micro-flow-regulator was tested.The double inlet spiral structure was verified by particle and cell experiments for cell sorting and integration with micro flow regulator;the integrated micro-chip containing micro-flow-regulator and double inlet spiral channel were studied.The functions of flow matching and flow stabilization were verified.The feasibility of commercial diaphragm pumps for cell separation by inertial microfluidic technology was verified.The flow rate of the sample solution and sheath solution at the two inlets of the spiral channel chip is maintained at 202±2μL/min and 920±5μL/min.The recovery rate of the two types of circulating tumor cells after sorting is 85.7%and 83.0%.