Development Of Cell Analysis Platforms Based On DNA-mediated Cell Surface Engineering And Microfluidic Chips

Cells are basic structural and functional units of living organisms.Understanding the composition,structure and function of cells,and exploring cellular activities,are quite important for the cognition of activity rules and phenomena of life.Cells have small size,low amount of contents and diverse species.And in organisms,cells are located in complex microenvironments and subjected to multiple cues that vary in time and space,including physical and biochemical factors,mechanical forces,and interactions with extracellular matrix(ECM)and other cells.Therefore,it is highly important and desirable to develop advanced technologies that enable precise cell manipulation,physiologically relevant microenvironment simulation,as well as high sensitive,selective,and reliable cell analysis.In this thesis,based on the DNA-mediated cell surface engineering strategy,and combined with mass spectrometry analysis and microfluidic techniques,we developed novel methods and platforms that enabled in-situ analysis of cellular components,realtime monitoring of cellular physiological processes and investigation of cell-cell interaction.First,we discussed the current states and major challenges in cell researches,and reviewed the important roles and recent advances of nucleic acid techniques,mass spectrometry analysis and microfluidic technology in the cell-related researches.Then,we developed a novel approach using DNA-mediated cell surface engineering for glycan profiling by MALDI-TOF mass spectrometry(MS).This strategy converted the analysis of glycans to the detection of DNA probes,overcoming the complicated composition and low ionization efficiency of glycans,enabling in situ detection and facilitating multiplex analysis.The amplification procedure also improved the sensitivity.This approach had been applied to evaluate glycomic alterations in cancer cells and provided the intrinsic distribution of glycans in tissues using MALDI imaging mass spectrometry.Next,based on the DNA-mediated cell surface engineering strategy,we designed a hairpin-type DNA sensor,which was randomly anchored on the cell membrane proteins and acted as an indicator for the cell endocytosis process.Combined with microfluidic devices,this DNA sensor was applied to investigate cellular mechanotransduction under fluid shear stress.Results revealed that cellular endocytosis process was significantly enhanced under shear stress,and this phenomenon was related to the specific endocytosis pathways.Finally,we developed a microfluidic device to study the cell-cell interactions in glioma angiogenesis.Glioma cells and endothelial cells were co-cultured in the microchannels,and the signal molecules secreted by glioma cells could induce the migration of endothelial cells.On the surface of endothelial cells,a specific aptamer sensor was anchored,which could respond to the corresponding signal molecule.Thus we could utilize this platform to real-time monitor the local concentration fluctuations of signal molecules,and correspond the changes to the cellular behaviors.