Western blots were performed as previously described (Kim et al. 2012 (link)). Protein concentration of cell lysates was determined by BCA protein assay (Thermo Fisher). Equal amounts of total protein for each lysate were separated on 4–20 % TGX™ (Tris-Glycine eXtended) gels (Bio-Rad) and transferred to 0.2 μm pore size nitrocellulose membranes (Bio-Rad). For Aβ detection, same volumes of media were run on 16.5% Tris-Tricine gel (Bio-Rad). After transferring, membranes were boiled for 10 minutes in PBS, as previously reported (McGowan et al. 2005 (link)). All membranes were blocked with 4% non-fat dry milk in TBS/T (Tris buffered saline with 0.125% Tween-20). Blots were probed with rabbit anti-APP antibody (Invitrogen), rabbit anti-BACE1 antibody (D10E5, Cell Signaling Technology), mouse anti-Aβ antibody (82E1, IBL International), rabbit anti-sAPPβ antibody (IBL International), rabbit anti-GAPDH antibody (FL-335, Santa Cruz Biotechnology), or mouse anti-actin antibody (Sigma) at room temperature for 1 h. Anti-APP antibody (Invitrogen) was used for CTFα and CTFβ detection. 6E10 (Covance) antibody was used for sAPPα detection. After secondary antibody incubation, membranes were developed using Lumigen TMA-6 ECL detection kit (Lumigen).