Ab237751

The TG tissues were removed from experimental mice on day 5 after corneal epithelial debridement, embedded in optimal cutting temperature compound, and stored at − 80 °C. Frozen sections were cut at a thickness of 5 μm. After fixing the sections with 4% paraformaldehyde, the sections were permeabilized and blocked, then incubated with the corresponding primary antibodies and Alexa Fluor-labeled secondary antibodies. Next, the tissues were counterstained with DAPI to visualize the cell nuclei. Finally, the fluorescence intensities were estimated using a fluorescence microscope (Leica, Germany). At least three independent experiments were performed. The primary antibodies used were as follows: anti-p62 antibody (P0067; Sigma-Aldrich, USA), anti-LC3B (ab192890; Abcam, USA), and anti-ATG4D (ab237751; Abcam). The imaging parameters of the same protein were consistent for all experiments (Additional file 1: Table S1).

The TG tissues were harvested on day 5 after corneal epithelial debridement and lysed with RIPA reagent (Beyotime, Shanghai, China) supplemented with 1% Phenylmethylsulfonyl Fluoride (PMSF) reagent (Beyotime). Protein samples were electrophoresed on Sodium dodecyl sulfate–polyacrylamide gels, transferred to polyvinylidene fluoride membranes, and blocked with skim milk. Afterward, the membranes were incubated in solutions of the corresponding primary antibodies and horseradish peroxidase-conjugated secondary antibodies. Eventually, Enhanced chemiluminescence was used to visualize proteins bands. At least three independent experiments were performed. The primary antibodies used for the study were as follows: anti-p62 antibody (p0067; Sigma-Aldrich, USA), anti-LC3B (ab192890; Abcam, USA), anti-ATG4D (ab237751; Abcam, USA), and anti-β-actin (ab8226; Abcam, USA).