High-throughput single-cell gene-expression profiling with multiplexed error-robust fluorescence in situ hybridization

We report advances that lead to a drastic increase in the measurement throughput of multiplexed error-robust fluorescence in situ hybridization, an image-based approach to single-cell transcriptomics

Jeffrey R. Moffitt


Scholarcy highlights

  • Image-based approaches to single-cell transcriptomics, in which RNA species are identified and counted in situ via imaging, have emerged as a powerful complement to single-cell methods based on RNA sequencing of dissociated cells
  • In most approaches to single-cell transcriptomics, cells are dissociated from tissues, and RNAs are extracted from cells; as a result, the native spatial context of these RNAs is lost
  • We report advances in multiplexed error-robust fluorescence in situ hybridization that increase the measurement throughput by two orders of magnitude and allow gene expression profiling of ∼40,000 human cells in a single 18-h measurement
  • The total time required for a MERFISH measurement can be divided into an area-dependent time that scales with the total imaged area and an area-independent time that does not
  • We found that the average copy number per cell for these 10 RNAs determined with MERFISH correlated strongly with the values determined via single-molecule fluorescence in situ hybridization
  • We describe several advances in the MERFISH method that increase the throughput of this approach by two orders of magnitude: We profiled 130 RNAs across 40 mm2 of sample containing as many as 39,000 human cells
  • We anticipate that with further optimization of the hybridization protocol, utilization of faster fluorescence signal removal protocols, incorporation of more colors per imaging round, and additional improvements in camera, optics, and light sources to increase the field of view area and reduce the imaging time further, it will be possible to increase the throughput of multiplexed error-robust fluorescence in situ hybridization further and to characterize millions of individual cells in their native culture and tissue contexts

Need more features? Save interactive summary cards to your Scholarcy Library.