Fig. 3.

Bronchiolar and alveolar architecture and histology of lung samples of control cases. Schematic illustrations (a,g; created with BioRender.com), brightfield images of H&E stained sections (b,h), and confocal images of sections (c-f,i) of control 1 and 2 stained with multiplex fluorescence RNAscope and IHC and DAPI as nuclear stain. (a) Schematic illustration of a transverse section through a bronchiole. (b–f) Images of sections through a bronchiole. (b) Brightfield H&E image of the bronchiolar wall. (c) FOXJ1 is a marker for ciliated cells in the bronchiolar epithelium and SFTPC is a marker for type 2 pneumocytes in the alveoli surrounding a bronchiole. The dashed square in (c) corresponds to the brightfield image of the same bronchiolar wall in a nearby section (b). In the bronchiolar epithelium, KRT5/6 is a marker for basal cells (d), CC16 for club cells (e) and MUC5AC for goblet cells (f). As expected, there are no N puncta (d–f). (g) Schematic illustration of a section through alveoli of the lung parenchyma. (h,i) Images of sections through alveoli. (h) Brightfield H&E image of alveoli. (i) KRT8 is a marker for type 1 and type 2 pneumocytes lining the alveoli and CD31 is a marker for endothelial cells lining the lumen of capillaries coursing through the alveolar walls; the weak orange-brown signal in the alveolar walls represents autofluorescence; no N puncta are detected. The names of RNAscope probes are in italics and the names of antigens/proteins that are revealed as immunoreactive signal by antibodies are in roman.