Enhanced chemiluminescence ecl kit

Chelerythrine, dithiothreitol (DTT) and 2,3,5-Triphenyltetrazolium chloride (TTC) were bought from Sigma Chemical Company (St Louis, MO). Antibodies against GRP78, CHOP, and caspase-12 were obtained from Abcam PLC (Cambridge, UK). Nitrocellulose membrane was obtained from Hybond-C (Amersham Life Science, UK) and the enhanced chemiluminescence (ECL) kit was provided by Beijing Applygen Technologies.

Protein concentrations were determined using a BCA Kit (Beyotime, Beyotime Biotechnology, Shanghai, China). Equal amounts of proteins were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis and transferred to polyvinylidene fluoride membranes. Antibodies against GPR41 (66811–1-Ig, Sanying, WuHan, China) PI3K (A4992, ABclonal, Wuhan, China), phosphorylated (p)-PI3K (Ab278545, Abcam, UK), Akt (A18675, ABclonal, Wuhan, China), p-Akt (AP1208, ABclonal, Wuhan, China), mTOR (A11355, ABclonal, Wuhan, China), p-mTOR (AP0115, ABclonal, Wuhan, China), RxRα (A19105, ABclonal, Wuhan, China), SREBP1 (A5754, ABclonal, Wuhan, China), PPARγ (A16958, ABclonal, Wuhan, China), St6gal1 (A5754, ABclonal, Wuhan, China), and GAPDH (AB-P-R001, Xianzhi, Hangzhou, China) were used as primary antibodies. All primary antibodies were used at a 1:1000 dilution except for those against p-PI3K (1:500 dilution) and GPR41 (1:2000 dilution). Goat anti-rabbit-HRP (BA1054, Boster Bio, Wuhan, China) and goat anti-mouse-HRP (BA1051, Boster Bio, Wuhan, China) were used as secondary antibodies. The bands were visualized using an Enhanced Chemiluminescence (ECL) Kit (Applygen, Applygen Technologies Co., Ltd., Beijing, China). The protein bands were densitometrically analyzed using the IPP software.