Ab71794

After sample processing, total protein was quantified using a bicinchoninic reagent kit (Thermo Fisher Scientific, Waltham, MA, USA). Samples were loaded at 30 μg protein volume with a 5 μL of a protein marker, and the protein concentration condition was set at 90 V for 30 min and the protein separation condition at 120 V for 50 min, depending on the protein size. The membrane was then transferred, washed with TBST (50 mL TBS + 450 mL ultrapure water + 250 μL Tween 20), and incubated with an FGF10 antibody (ab71794, RRID: AB_2049648, Abcam, Cambridge, UK) overnight at 4 °C. After washing with TBST, the membranes were then incubated with the secondary antibody, washed for 1 h, and then visualised for protein quantification.

Lung tissues were homogenized, and total proteins were extracted using tissue lysis buffer supplemented with phenylmethanesulfonyl fluoride. In the in vitro experiment, the total proteins of AECs were extracted using a cell lysis buffer. For the quantification of protein content, a BCA kit (Beyotime, P0010S) was used. Denatured proteins were added to denaturing gels and separated by SDS-PAGE (Mini-PROTEAN Tetra electrophoresis system, Bio-Rad). The separated proteins were then transferred onto polyvinylidene fluoride membranes, and the membranes were incubated with primary antibodies. Primary antibodies against LC3B (Abcam, ab128025,1:1000), P62 (Abcam, ab56416,1:1000), BMP4 (Abcam, ab39973,1:800), SPC (Abcam, ab90716,1:900), FGF10 (Abcam, ab71794,1:1000), FGFR2 (Abcam, ab10648,1:2000) and GAPDH (Bioworld, AP0063,1:20000) were used at 4°C overnight after blocking with skim milk. The PVDF membranes were incubated with secondary antibodies (BioWorld, goat anti-rabbit, BS13278; goat anti-mouse, BS12478) and visualized by chemiluminescence the following day. Finally, the gray value of each band was measured using ImageJ software.