Ab264305

Western blot analyses were performed as previously described (Lin et al., 2018) (link). Briefly, cells were washed 3 times with PBS and were lysed at 4°C with RIPA buffer (P0013C, Beyotime Biotechnology) containing phenylmethylsulfonyl fluoride (0.5 mM, ST506, Beyotime Biotechnology), aprotinin (5 μg/mL, A1153, Sigma-Aldrich), and leupeptin (5 μg/mL, L2884, Sigma-Aldrich). Approximately 35 μg of proteins was separated on 10% SDS-PAGE and blotted onto nitrocellulose membrane. Membranes were blocked for 1 h at room temperature with 5% skim milk in Tris-buffered saline with 0.05% Tween 20 and incubated overnight at 4°C with primary antibodies at the following dilutions: CIDEC (1:1,000 dilution; ab198204, Abcam), FASN (1:1,000 dilution; Cell Signaling Technology), C/EBPβ (1:1,000 dilution; ab264305, Abcam), and β-actin (1:1,000 dilution; 4970, Cell Signaling Technology). The blots were then incubated with the appropriated secondary antibodies conjugated to horseradish peroxidase (1:500 dilution; ZSGB-Bio) at room temperature for 1 h. Proteins were visualized using enhanced chemiluminescent detection reagent (6683, Signaling Technology). Densitometric analysis was performed with Image-Pro Plus 6.0 (Media Cybernetics Inc.), and target protein expression was normalized to β-actin expression.