Practical Considerations for Complex Tissue Dissociation for Single-Cell Transcriptomics.

ABSTRACT

Single-cell and single-nucleus RNA sequencing have revolutionized biomedical research, allowing analysis of complex tissues, identification of novel cell types, and mapping of development as well as disease states. Successful application of this technology critically relies on the dissociation of solid organs and tissues into high-quality single-cell (or nuclei) suspensions.In this chapter, we examine several key aspects of the tissue handling workflow that need to be considered when establishing an efficient tissue processing protocol for single-cell RNA sequencing (scRNA-seq). These include tissue collection, transport, and storage, as well as the choice of the dissociation conditions. We emphasize the importance of the tissue quality check and discuss the advantages (and potential limitations) of tissue cryopreservation. We provide practical tips and considerations on each of the steps of the processing workflow, and comment on how to maximize cell viability and integrity, which are critical for obtaining high-quality single-cell transcriptomic data.