Sc 49437

Immunoblots were performed as previously described [5 (link)]. Antibodies against p16 (1:200 dilution; #sc-1207), ALDH2 (1:200 dilution; #sc-100496) and acetylated FoxO1 (1:200 dilution; #sc-49437) were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Antibodies against p53 (1:1000 dilution; #2524), LC3B (1:1000 dilution; #3868), SirT1 (1:1000 dilution; #3931), acetyl-lysine (1:1000 dilution; #9441), Atg7 (1:1000 dilution; #8558), Rab7 (1:1000 dilution; #9367) and FoxO1 (1:1000 dilution; #2880) were purchased from Cell Signaling Technology. Antibody against DNPH (1:2000 dilution; #ab93160), LAMP2 (1:1000 dilution; #ab25339), Tubulin (1:1000 dilution; #ab179513), GAPDH (1:10000 dilution; #ab181602) and TBP (1:5000 dilution; #ab28175) were purchased from Abcam. Antibody binding was detected via enhanced chemiluminescence (Millipore) and scanned with ChemiDocXRS (Bio-Rad Laboratory, Hercules, CA). Immunoblot band intensity was analyzed with Lab Image software. For immunoprecipitation analysis, lystes were mixed with primary antibody. Immunocomplexes were separated by SDS-PAGE and detected with western blot analyses.

Protein levels of ac-NF-кB, ac-FoxO1, SIRT1, and ac-p53 in brain samples were evaluated by western blotting according to standard protocols. In brief, the protein samples were blotted onto the PVDF membrane. Then, the membrane was incubated with primary antibodies against SIRT1 (Ab12193, 1 : 1000, Abcam), ac-FoxO1 (sc-49437, 1 : 200, Santa Cruz), ac-NF-кB (12629S, 1 : 500, Cell Signaling), ac-p53 (2570, 1 : 500, Cell Signaling), and appropriate secondary antibodies. β-Actin (BM0627, 1 : 4000, Boster Biotech) was the internal loading control for the proteins. Membranes were observed with enhanced chemiluminescence solution.