μ dish 35 mm imaging dishes

LS174T cells were seeded on μ-dish (35-mm) imaging dishes (ibidi GmbH, Martinsried, Germany) and treated with 30 mM succinate for 12 h. Then, the cells were fixed in 4% paraformaldehyde in PBS for 10 min, permeabilized with PBS containing 0.1% Triton X-100 for 5 min, and incubated with a primary antibody against MUC2 as (ab97386) for 24 h at 37°C. The cells were then incubated with rhodamine-phalloidin and Hoechst 33352 to stain the F-actin and nuclear chromatin, respectively. Thereafter, the cells were examined using an All-In-One Fluorescence Microscope BZ-X810 laser scanning microscope (Keyence, United Kingdom).

LS174T cells were cultured in a 24-well-plate or μ-dish (35-mm) imaging dishes (Ibidi GmbH, Martinsried, Germany) until they reached 100% confluence.
Then, cells were treated with 10 μM AKT inhibitor (perifosine) for 1 h before succinate treatment. Acetate (FUJIFILM Wako Pure Chemical Co., Ltd.), butyrate (Nu-Check Prep, Inc, Waterville, MN, USA), propionate (Nu-Check Prep, Inc, Waterville, MN, USA), and succinate (Nu-Check Prep, Inc, Waterville, MN, USA) were dissolved in phosphate-buffered saline (PBS) in this experiment.