Ab66613

Dissected human or mouse liver tissues were fixed in 4% paraformaldehyde at 4 °C for at least 12 h and incubated in 30% sucrose at 4 °C overnight. Samples were embedded in the optimum cutting temperature (OCT) compound and cut into 5-μm sections. After blocking with TBST containing 20% FBS, sections were incubated with anti-FXYD1 (Abcam, ab76597, 1:500), anti-GJB1 (Abcam, ab66613, 1:500), anti-HNF4A (Santa Cruz Biotechnology, sc-6556, 1:50), anti-SOX9 (Millipore, ab5535, 1:200), anti-FGB (Abcam, ab118533, 1:500), anti-ID3 (Abcam, ab41834, 1:500), and anti-VEGFR3 (Invitrogen, 14-5988-82, 1:100) antibodies at 4 °C overnight. After washing, sections were treated with Alexa Fluor 488 donkey anti-goat IgG (Invitrogen, A11055, 1:1000), Alexa Fluor 594 donkey anti-goat IgG (Invitrogen, A11058, 1:1000), Alexa Fluor 488 donkey anti-rabbit IgG (Invitrogen, A11008, 1:1000), Alexa Fluor 594 donkey anti-rabbit IgG (Invitrogen, A21207, 1:1000), Alexa Fluor 647 donkey anti-rat IgG (Jackson ImmunoResearch, 712-605-150, 1:250), and Alexa Fluor 488 donkey anti-sheep IgG (Invitrogen, A11015, 1:1000). DAPI (Sigma, D9564, 0.5 µg/mL) was used for nuclear staining. Images were acquired using an LSM 710 NLO and DuoScan System (Zeiss).

The SH-SY5Y cells (2 × 10⁵ cells/well) were seeded onto a 3 cm-diameter culture dish and cultured 24 hours. Immunofluorescent staining was conducted in accordance with our previous studies [55 (link)]. Primary Abs used were Cx32 (Abcam, ab66613), Nur77, LC3 (Santa Cruz Biotechnology, sc365113, sc271625), Rac1 (Movus, NBP2-67091), β-tubulin (CST4466).