Immunofluorescence of Liver Tissue Sections

Immunofluorescence was performed, as described in detail recently without modifications^{6 (link)}. Specifically, liver tissue was fixed with paraformaldehyde, cryo-protected in sucrose, and frozen in O.C.T. (Tissue-Tek). Frozen liver sections (5 μm) were permeabilized in ice-cold methanol, then room temperature 0.1% Triton X-100, and then reacted with DAPI at 0.08 ng/mL (Invitrogen, D1306) and the antibodies indicated below. Anti-eGFP (Invitrogen, #A-11122), Anti-HNF4α (Abcam, #ab41898), Anti-HAL (Sigma Prestige Antibodies, #HPA038547), anti-Cytokeratin 7 (Abcam, #ab68459), anti-albumin (Bethyl, #A80229A), anti-KIAA0319L (AAVR, Abcam, #ab105385), anti-CD68 (Invitrogen, #14068882) antibodies were used as indicated. For Figs. 3k, and 4h–k antigen retrieval was required. This was achieved by heating the slides for ten minutes in antigen retrieval buffer (Tris-EDTA, pH 9). After immunolabelling, the images were captured and analyzed on a LSM800-Airyscan microscope using ZEN Black software.