Laminin

Cells or tissue sections were fixed, permeabilized, and incubated with primary antibodies [Iba1 (wako), laminin (Boster), CD31 (Boster, Abcam, HuaBio), albumin (Invitrigen), YFP (SAB), ZO‐1 (Proteintech), mCherry (Abcam), MMP9 (Boster), Aqp4 (Abcam), PDGFRβ (Abcam)] overnight at 4 °C, followed by incubation with TRITC‐conjugated, Alexa647‐conjugated, or FITC‐conjugated secondary antibody (Jackson ImmunoResearch) and counterstained with Hoechst33342 (Sigma‐Aldrich). Fluorescence images were captured using fluorescence microscopy (DXM1200, Nikon) or confocal microscope (TCS SP5, Leica) and quantified using Image‐Pro Plus (Media Cybernetics) or Image J (NIH Image, Bethesda, MD).^[45]

On day 5 after recovery, the retinal cups were fixed with 4% paraformaldehyde for 30 min, sucrose gradient dehydration and embedded in optimal cutting temperature (OCT) compound overnight at − 80 °C. The cups were sliced at 10 μm using a Leica microtome. After being permeabilized with 0.5% Triton X-100 (Sigma-Aldrich, St Louis, MO, USA) and blocked with normal goat serum (Boster, Wuhan, China), sections were stained with primary antibodies against Ki67 (Abclonal, USA), NEFL (Abclonal, USA), Laminin (Boster, Wuhan, China), and cleaved caspase-3 (CST, MA, USA) Brn3 (Santa Cruz Biotechnology, Dallas, TX) HuC/D (Santa Cruz Biotechnology, Dallas, TX) Recoverin (Abcam,Cambridge, U.K.) OTX2 (Abcam, Cambridge, U.K.) AP2α (Developmental Studies Hybridoma Bank (DSHB), Iowa, USA) PAX6 (Developmental Studies Hybridoma Bank (DSHB), Iowa, USA) at 4 °C overnight. The secondary antibodies used included the corresponding species-specific Alexa Fluor-555-, and Alexa Fluor-647-conjugated antibodies (1:500; Gibco, Carlsbad, CA) and nuclei were stained with 4′,6-diamidino-2-phenylindole (DAPI). Photos were captured with a fluorescence system (BX50; Olympus, Tokyo, Japan).