Microfluidic Single Cell Metabolite Analysis

Typical microfluidic devices have dimensions of tens to hundreds of micrometers which are comparable to the size of a single cell,therefore,the microfluidic devices are quite suitable for single-cell analysis.Among various microfluidic techniques,droplet-based microfluidics is capable of generating droplets containing single cells by an oil phase where the volumes of droplets are typically in the range of nanoliters to picoliters.The immiscible oil surrounded the droplets could eliminate sample adsorption on the solid surface,sample dilution and cross contamination.The combination of droplet microfluidics and capillary electrophoresis with laser induced fluorescence detection(CE-LIF)provides a new platform for metabolite analysis in single cells.In chapter 1,the technologies based on microfluidics for single-cell metabolite analysis which were divided into two major categories,namely in-situ analysis and separation-based analysis were reviewed.According to the different detection methods,the two kinds of technology were further introduced in details.In chapter 2,a chiral separation system for cell metabolite analysis was developed based on the combination of microfluidic nanoliter droplet array and CE-LIF.The conditions for aspartate(Asp)derivation reaction and the chiral separation of Asp were optimized,including the composition and concentration of separation buffer,the injection time,the category of internal standard,the concentration and solvent of NDA,the concentration of NaCN,and the pH of the derivatization reaction.The chiral analysis system was further applied to the detection of Asp in rat adrenal pheochromocytoma cells and the single cell in the cerebral ganglion of Aplysia California.The influences of the culturing time of cells,the condition of digestion and the method of lysis were investigated.The extra influences of the ways of droplet generation and reagents addition were investigated in the cerebral ganglion of Aplysia California.The quantitative analysis of D-Asp in single cells in the cerebral ganglion of Aplysia California was achieved.