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 the 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, 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.

Nanodroplet-Based Reagent Delivery into Water-in-Fluorinated-Oil Droplets

“…Semi-permeable hydrogels [ 28 ] or microcapsules [ 29 ] are another state-of-the-art approach for controlling the biochemical reactions within micro-compartments. In these systems, the different reagents smaller than the pore size can diffuse into micro-compartments.…”

“…The peptide concentration increased only slightly to 34 nM when the incubation was continued for a total of 24 h (Figure S3), which suggested that the peptide delivery was mostly completed in the first 3 h. These results demonstrated the feasibility of using nanodroplets to achieve delivery of medium-sized biomolecules into water-in-fluorinated-oil microdroplets. Semi-permeable hydrogels [28] or microcapsules [29] are another state-of-the-art approach for controlling the biochemical reactions within micro-compartments. In these systems, the different reagents smaller than the pore size can diffuse into micro-compartments.…”

Lab on a Particle Technologies

“…The first applications of hollow microparticles have been the selection of cells based on growth, RNA cytometry, drug delivery, cell delivery, , water treatment, and enzymatic analysis . Atrandi Biosciences has commercialized instruments and microfluidic kits for sequencing microbes using hollow shell particles .…”

Harnessing HetHydrogel: A Universal Platform to Dropletize Single‐Cell Multiomics

“…Researchers have devised various compartmentalization methods, such as microwells or homogeneous polymer beads, to enable an open yet isolated system for massively parallel biochemical reactions. [11][12][13] Hydrogel, a biocompatible and inherently porous material, has been extensively used to house biomolecules and cells. [14,15] Hydrogel-based compartments create a permeable environment, enabling the necessary substance exchange for biochemical reactions and cell growth.…”