Protocol to dissociate healthy and infected murine- and hamster-derived lung tissue for single-cell transcriptome analysis

In infectious disease research, single-cell RNA sequencing allows dissection of host-pathogen interactions. As a prerequisite, we provide a protocol to transform solid and complex organs such as lungs into representative diverse, viable single-cell suspensions. Our protocol describes performance of vascular perfusion, pneumonectomy, enzymatic digestion, and mechanical dissociation of lung tissue, as well as red blood cell lysis and counting of isolated cells. A challenge remains, however, to further increase the proportion of pulmonary endothelial cells without compromising on viability.