A Microfluidic Chip For Studying Single-cell Signal Transduction At High Time Resolution

Microfluidic chip has become an important way to study single cell because of its unique structure.It has the advantages of high precision,low cost,few samples required,fast reaction speed and easy parallelization experiment.In order to make the research method more convenient,the chip that can capture,culture and analyze the single cell has become a research hotspot.In this paper,single drop single cell microchip system is used as the research platform,single cell capture,analysis,droplet stimulation and rapid detection are integrated,and with the intersection of immunocytochemical methods,the rapid dynamic process of protein phosphorylation in single cell under high time resolution is realized,providing a new technology and new method for the study of single cell signal transduction pathway and molecular mechanism.In this paper,we develop a new analysis system based on surface energy for single drop single cell microchips,which can analyze the rapid kinetic process of protein phosphorylation in situ with high time resolution,and realize the study of the rapid signal transduction pathway of single cell.Specific research contents include:(1)A microfluidic chip containing a dual inlet and Surface Energy Well(SEW)array was prepared by simply changing the flow velocity of the continuous phase.(2)The experimental results show that the replacement time of droplet is as fast as 3 seconds,and the reagent exchange rate reaches 100%,which indicates that the reagent can be completely replaced by single droplet replacement,and the initial stimulus can be completely stopped.(3)Adherent cells were cultured at the bottom of microchannel on a chip under continuous culture solution perfusion,and single cell culture was achieved by utilizing fluid dynamics flow changes in the SEW region.(4)The PDGF/Akt pathway was selected as a model system for rapid cellular events to monitor the kinetics of receptor phosphorylation in response to cellular stimulation at a temporal resolution of seconds.Therefore,we suggest that this approach could potentially be used as an important new strategy for a wide range of studies to detect the temporal dynamics of fast cellular events at the single-cell level.The method provides a common platform for probing the temporal dynamics of single cells and for monitoring rapid chemical reactions in a variety of applications.