Highly multiplexed quantitation of gene expression on single cells

Abstract

Highly multiplexed, single-cell technologies reveal important heterogeneity within cell populations. Recently, technologies to simultaneously measure expression of 96 (or more) genes from a single cell have been developed for immunologic monitoring. Here, we report a rigorous, optimized, quantitative methodology for using this technology. Specifically: we describe a unique primer/probe qualification method necessary for quantitative results; we show that primers do not compete in highly multiplexed amplifications; we define the limit of detection for this assay as a single mRNA transcript; and, we show that the technical reproducibility of the system is very high. We illustrate two disparate applications of the platform: a “bulk” approach that measures expression patterns from 100 cells at a time in high throughput to define gene signatures, and a single-cell approach to define the coordinate expression patterns of multiple genes and reveal unique subsets of cells.

Single cells from human primary colorectal tumors exhibit polyfunctional heterogeneity in secretions of ELR+ CXC chemokines

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Single-cell technologies for monitoring immune systems

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“…In particular, reagent panels must be carefully constructed, to ensure that the quality and utility of data is maximized 55,61–63 . To ease this process, qualified reagents can be grouped into ‘cassettes’ based on the shared or complementary biology of the markers they detect.…”

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