Pan ck

Samples were fixed in 4% paraformaldehyde in phosphate buffered saline (PBS) and then embedded in paraffin using standard procedures. Serial paraffin sections (4-μm thickness) were prepared for Hematoxylin and Eosin (HE), immunostaining and in situ hybridization. Antigen retrieval was achieved by citrate buffer, pH 6.0. After antigen retrieval, immunohistochemical analyses were performed using the DakoCytomation Envision System (using horseradish peroxidase with diaminobenzidine enhancer) (Dako, United States) according to the manufacturer’s instructions. The slides were incubated with antibodies against Pan-cytokeratin (Pan-CK) (1:50, Thermo Fisher Scientific^TM, United States), FGF10 (1:50, Santa Cruz, Unite States), Caspase 3 (1:1600, Cell Signaling Technology, United States) and Ki67 (1:200, Abcam, United Kingdom). The specimens were sequentially incubated with secondary antibody and streptavidin peroxidase. Finally, the results were visualized following staining using a diaminobenzidine reagent kit (Invitrogen, United States). The sections were counterstained with hematoxylin. All specimens were observed by stereomicroscope (MD5500D; Leica, camera: DFC495; Leica, Lens: HCX PL APO 409; Leica). At least 10 mice were examined in each experiment.

Mouse lung tissue was embedded in paraffin, sectioned and stained with hematoxylin and eosin. Immunofluorescence staining was performed to detect T1alpha (Developmental Studies Hybridoma Bank clone 8.1.1), SPC (Millipore/Chemicon, AB3786), K14 (made at the Antibody Facility in NIBS), CD31 (BD Biosciences, 553369), vimentin (DAKO, IR63061), CK8 (Maixin, MAB-0167), Pan-CK (Thermo Scientific, MA5-13203), and SMA (Sigma, c6198).