RNA cytometry of single-cells using semi-permeable microcapsules

Abstract

Analytical tools for gene expression profiling of individual cells are critical for studying complex biological systems. However, the techniques enabling rapid measurements of gene expression on thousands of single-cells are lacking. Here, we report a high-throughput RNA cytometry for digital profiling of single-cells isolated in liquid droplets enveloped by a thin semi-permeable membrane (microcapsules). Due to selective permeability of the membrane, the desirable enzymes and reagents can be loaded, or replaced, in the microcapsule at any given step by simply changing the reaction buffer in which the microcapsules are dispersed. Therefore, complex molecular biology workflows can be readily adapted to conduct nucleic acid analysis on encapsulated mammalian cells, bacteria, or other biological species. The microcapsules support sequential multi-step enzymatic reactions, and remain intact under different biochemical conditions, freezing, thawing and thermocycling. Combining microcapsules with conventional FACS provides a high-throughput approach for conducting RNA cytometry of individual cells based on their digital gene expression signature.