Ab7818

Western blot analysis was performed as described previously.²² (link) Primary antibodies were anti‐IL‐22 (ab181007; Abcam), anti‐TGF‐β1 (ab92486; Abcam), anti‐α‐SMA (ab7818; Abcam), E‐cad (3195S; CST, USA), TGF‐βR2 (ab61213; Abcam), Vimentin (5741S), Smad2/3 (8685S) and P‐smad2/3 (8828S; CST), GADPH (14C10; CST), Vimentin (ab92547; Abcam), Fibronectin (FN) (610077; BD Systems, USA), Collagen‐I (ab34717; Abcam). Secondary antibodies were goat anti‐rabbit and rabbit anti‐mouse secondary antibodies (7076S and 7074S; CST).

Immunofluorescence were performed in TGF-β1-treated ESCs to detect the co-localization of GLS1 and α-SMA. ESCs were seeded on 35 mm confocal dishes (Cellvis) at a density of 1 × 10⁴ cells/mL. After treatments, cells were fixed with 4% PFA for 15 min, permeabilized with 0.2% Triton X for 10 min and then blocked with 5% BSA for 45 min at room temperature. Primary antibodies α-SMA (1:200; Abcam, ab7818), GLS1 (1:200; Abcam, ab93434) were used for overnight incubation at 4 °C. DyLight488-conjugated goat anti-rabbit antibody (1:200; Multisciences, GAR4882) and DyLight594-conjugated goat anti-mouse antibody (1:200; Multisciences, GAM5942) were used for detection. DAPI was used to stain cell nucleus, and the stained ESCs were observed by an Olympus FV1200 confocal microscope (Olympus).