Single Cell RNA-Seq: Cell Isolation and Data Analysis

Single-cell RNA-sequencing technologies have revolutionized the way that researchers can interrogate cellular relationships and the level of detail by which tissue architecture can be characterized. Multiple cell capturing methods have been developed that, when coupled to next-generation sequencing, can yield cell-to-cell specific information regarding gene expression profiles. One of the commonalities between all of the cell capturing techniques to succeed is the necessity to submit samples with a high cell viability. In addition, these cells should have undergone minimal processing to limit induced stress responses so that their transcriptomes, when sequenced, closely reflect their transcriptomes in vivo at the time of isolation. Below we present a streamlined protocol to isolate fresh cells from tissues in vivo. We also share extensive notes to highlight considerations researchers should take into account before beginning their cell isolation protocol.