Study On The Capture Of Human Peripheral Blood Lymphocyte Based On Microfluidic Chip

The detection of lymphocytes in peripheral blood is a reliable method to evaluate the influence of disease on human body in the early time.Core/shell Fe₃O₄@Au magnetic nanoparticles（MNPs）are the attracted increasing attention in the field of nano biology sense such as the capture of bacteria,cells and DNA due to their prominent magnetic enrichment.Besides,solid structures prepared by mental deposition were also utilized as the tools to capture the lymphocytes.Microfluidic chip is a measurement platform with high flux and micromation,which can be applied in the capture and detection in cells in a controllable and repeatable method.With the advantages of less sample,simple operation and high accuracy,this technique has a very broad application in the detection of lymphocytes.The main work is as follows:（1）The synthesis of core/shell Fe₃O₄@Au magnetic nanoparticles are summarized.The superparamagnetic microspheres（400⁶00nm）were synthesized by the method of solvent thermal and the Fe₃O₄@Au magnetic nanoparticles were prepared by the method named sonochemically assisted with hydroxylamine seeding growth.An detailed summary of the synthesis of Fe₃O₄@Au magnetic nanoparticles and the synthesis method for Fe₃O₄@Au magnetic nanoparticles based on the published references are mentioned.The controllable preparation of magnetic microspheres（400⁶00 nm）were successfully completed with exploration in the experiments through the method of solvent thermal,which established substantial grounding.The Fe3O4@Au magnetic nanoparticles with great equality in size and form were synthesized through the method named sonochemically assisted with hydroxylamine seeding growth,which was mentioned by our research group previously.TEM,SEM,EDS,XRD,UV-vis and magnetormeter were implemented to characterize the synthesized products.In addition,the synthesized Fe₃O₄@Au MNPs have a good SERS performance.According to the practical requirement,different thickness and complete continuity Au shell was grown on the magnetic core.Compared with the traditional methods,the method mentioned above has the advantages like rapid reaction time,good cladding and easy operation.The solid structure with 200nm Au film was fabricated by mental deposition（2）Lymphocytes in peripheral blood were capture by two kind of structures synthesized previously according to the principle of antigen-antibody specific binding.Fe₃O₄@Au magnetic nanoparticles linked with anti-CD₃ specifically bind with CD₃on the surface of lymphocytes,then concentrated by magnetic field,which lymphocytes were separated and captured from the peripheral blood.First of all,the separated lymphocytes collected through density gradient equilibrium method were captured to prove the practicability.Then the condition of incubation time and the concentration of lymphocytes were discussed to optimize the efficiency.Finally the lymphocytes in peripheral blood were captured with the modified experimental parameters.An stable,high specific binding solid structure was reported to capture the lymphocytes in peripheral blood.With self-assembly,the Au film linked with anti-CD₃and the lymphocytes were then captured on the surface of Au film.The captured lymphocytes can be fixed in a certain area,which can quantization the number of lymphocytes.（3）The contents of fluid flow at the microscale and the synthesis of microfluidic chip are introduced.A kind of microfluidic,“Y”structure was designed for the capture and radiation damage detection of lymphocytes.First,the fluid flow of existing microfluidic chip was introduced.According to the theory of fluid mechanics,a kind of microfluidic,“Y”structure was designed.The SU-8ultraviolet photoresist mold was fabricated by spin-coating,photo-lithography and development.The technological parameter in the experiment of the micro was studied and the technological parameter was optimized.At the same time,the Fe3O4@Au magnetic nanoparticles was driven controllably in the microfluidic chip successfully.（4）A complete platform for the capture of lymphocytes was set up,the capture of lymphocytes was carried out successfully in the microfluidic chip.For the demand of capture of lymphocytes,a platform consists of microfluidic chip,micro-injector pump,Stereoscopic microscope was set up.First,the separated lymphocytes collected through density gradient equilibrium method was added into the microfluidic chip and the experimental parameters was modified to increase the efficiency of lymphocytes capture.The limit of lymphocytes capture was also studied.In addition,the microfluidic chip filled in solid structure was also carried out to realize the capture of lymphocytes.