Differential in vivo labeling with barcoded antibodies allows for simultaneous transcriptomic profiling of airway, lung tissue and intravascular immune cells

We used single-cell RNA sequencing (scRNA-seq) paired with DNA-barcoded antibodies to see if it allows to distinguish immune cells in different anatomical compartments in the lung akin to flow cytometry. To do this, we infected Balb/c animals with non-typeable Haemophilus influenzae (NTHi) and administered barcoded anti-CD45 antibody intratracheally and intravenously to label airway and vascular immune cells respectively. We isolated and sequenced neutrophils from the lung as these cells undergo transendothelial migration upon inflammation. We sequenced 5,543 neutrophils and found 3 different major clusters of neutrophils based on barcoded CD45 antibodies. Single-cell RNA sequencing with feature barcodes was performed on lung neutrophils from Balb/c animals infected with NTHi for 48-hours. For this experiment, cells were stained with 32 DNA-barcoded antibodies