UC Irvine

Single-cell analysis is of critical importance in revealing population heterogeneity, identifying minority sub-populations of interest, as well as discovering unique characteristics of individual cells. Conventional bench-top methods are limited by their high cost, low throughput, and inadequacy in analyzing small amount of material. Microfluidic platforms, on the contrary, work at the scale comparable to cell diameter, and recent advances in microfluidics have made it possible to automate the processing and analysis of single cells in a high-throughput and low-cost manner.

In this dissertation, a system of microfluidic platforms enabling high-throughput single-cell analysis from phenotype to genotype is developed. The system is built upon a microfluidic trapping array for rapid and deterministic single-cell trapping in highly-packed microwells. It is a serpentine channel with microwells arrayed along each row, wherein cells are delivered to the traps sequentially by the horizontal flow and pushed into traps by the perpendicular stream through the gap area at each trap. 1600 microwells are filled within 3 min with a single-cell occupying efficiency > 80%, while smaller cells/debris are filtered out simultaneously.

Two innovative single-cell analyses from phenotype to genotype are established on this single-cell array: live-cell real-time metabolic imaging via fluorescence-lifetime-imaging-microscopy (FLIM), and single-cell mRNA live-probing by dielectrophoretic nanotweezers (DENT). Rapid trapping and identification (both by FLIM and mRNA probing) of single circulating-tumor-cells (CTCs) from blood have been successfully demonstrated. After characterizing the individual trapped cells, in order to edit aberrant genes for the identified cells of interest, a droplet-microfluidic platform has been developed for efficient single-cell transfection (10X higher efficiency for suspension cells compared to the bulk approach). To explore cell-cell interaction at single-cell level, the single-cell array has also been modified into an easy-to-operate cell-pairing array.

As the trapping efficiency is determined by the channel parameter instead of the flow rate, the single-cell array can be integrated with various sample-processing units operating at different flow rates. The presented microfluidic system enables high-throughput single-cell trapping, label-free metabolic imaging, mRNA extraction without cell lysing, efficient gene transfection, and cell-cell interaction analysis. It is expected to have myriad applications in cancer diagnostics, gene therapy, immunology, etc.