An automated method to measure the expression of genes in thousands of single human cells is reported this week in Nature Methods. The method scales up a technique known as single-molecule fluorescence in situ hybridization (smFISH) and determines where RNA transcripts are located in the cell, providing important clues about their biological function.
smFISH can be used to detect specific RNA sequences in a cell based on their binding to a fluorescent probe, but imaging the ‘dots’ corresponding to each RNA molecule requires high magnification and delicate imaging settings. Lucas Pelkmans and colleagues use much brighter probes, allowing them to perform rapid and robust low-magnification imaging of many more cells, quantify low-level expression accurately and also query very short RNA transcripts. Their software tools automatically outline cells and nuclei, count dots to quantify expression and exhaustively document where transcripts are located in the cell. Results from studies with human cells showed highly reproducible expression levels, comparable with those determined by high-throughput RNA sequencing (RNA-seq).
The authors highlight the importance of measuring the variability of transcript expression and location in so many cells by using their data to discover genes with related functions in the cell.