Droplet-based Mcriofluidic Single Cell Analysis System

In rescent years, droplet-based microfluidics has made great progress and its application in single cell analysis has attracted more and more attension. Droplets, which are used as single cell microreactors, have the advantages of limiting dispersion, accelerating mixing and improving detection sensitivity. Despite the great success of droplet-based systems in single cell analysis, a multifunctional microfluidic platform that could perform complex single cell analysis with multiple manipulations of droplets including generation, transport, combining and splitting still present challenges.In Chapter One, the recent progress of microfluidic single cell analysis systems are reviewed, including the single phase microfluidic single cell analysis system and the droplet-based microfluidic single cell analysis system. In the latter section, various droplet manipulation techniques are comprehensively described including droplet generation, reagent introduction, droplet fusion and multi-step manipulation. The application of droplet microfluidics in single cell encapsulation, sorting, culture and analysis are also introduced.In Chapter Two, an automated doplet-based microfluidic single cell anlysis system based on a capillary sampling probe and a slotted-vials array was developed. The single cell droplet was formed using droplet assembling technique by sequentially aspirating cell suspension, reagent solutions and oil carrier into the capillary. Various factors affecting single cell encapsulating efficiencies were investigated. Under optimized conditions, a high single cell encapsulation efficiency of 46.7±2.0% was obtained, which was 10% higher than those reported by other groups. The performance of the system was demonstrated in the intracellularβ-galactosidase activity assays of pheochromocytoma cells (PC 12 cells) at the single cell level. With the ability in generating droplets with different compositions, simultaneous analysis for different cell samples was also performed. In Chapter Three, a novel and simple droplet fusion method was developed with the single cell analysis system described in Chapter Two, based on the use of the difference of droplet interfacial tensions. By varying the droplet interfacial tension, the velocity differences between the droplets was generated, and droplet fusion in a capillary-based system without any external devices was achieved. The droplet fusion techinique was applied in on-line monitoring of enzymatic reaction dynamic process of single cells in a short period.In Chapter Four, multi-step magnetic droplet manipulation technique was developed with the single cell analysis system described in Chapter Two. By introducing ferromagnetic particles as carriers to assist the magnetic beads in passing through the phase interface, picoliter-scale droplet splitting was achieved. By employing multi-step droplet manipulations including droplet transferring, merging and splitting, solid phase extraction among picoliter-scale droplets was performed and applied in DNA purification for single cell or small numbers of cells.