Anti actin

The cells were lysed in buffer containing 50 mM HEPES, 2.5 mM EDTA, 1 mM PMSF, 5 μg/mL aprotinin, and 10 μg/mL leupeptin; 30 μg of protein lysate were electrophoresed on 8 or 10% SDS-PAGE gels and transferred to methanol-activated PVDF membranes. The membranes were blocked with 5% non-fat skim milk and incubated with primary antibodies at 4 °C overnight. Western blotting was visualized by peroxidase-conjugated secondary antibodies and ECL chemiluminescent substrate (Immobilon Western Chemiluminescent Substrate, Millipore, Burlington, MA, USA). The quantification of target protein bands with reference to control bands (for each concentration) used the ImageJ Gel Analysis program.
The following primary antibodies were used: anti-Aβ1-40 (A-8326, Sigma-Aldrich), anti-Aβ1-16 (MAB 5208. Merck, Kenilworth, NJ, USA), anti-APP-CTF (AB5352, Merck), anti-IDE (AB9210, Merck), anti-NEP (AB5468, Merck), anti-LC3 (2775, Cell Signaling Technology, Danvers, MA, USA), anti-p62 (8025S, Cell Signaling Technology), anti-p-AMPK(Thr172) (2535S, Cell Signaling Technology), anti-AMPK (2532S, Cell Signaling Technology), anti-GAPDH (GTX100118, GeneTex, Hsinchu, Taiwan), and anti-actin (AC-15, Novus, Centennial, CO, USA). Secondary antibodies: HRP-linked anti-rabbit IgG (7074, Cell Signaling Technology) and HRP-linked anti-mouse IgG (Jackson ImmunoResearch Laboratories, West Grove, PA, USA).