Hrp conjugated rabbit of mouse secondary antibodies

Cell lysates were determined the protein concentration using the BCA assay (Pierce, Rockford, IL, USA) and 10 μg protein samples were separated on 8–16% gradient SDS-PAGE gels (Life technologies, Carlsbad, CA, USA) and transferred to nitrocellulose membrane (0.45 μm pore size, Bio-Rad). After 30 min incubation with 5% skim milk in TTBS buffer (10 mM Tris (pH7.5), 150 mM NaCl, 0.05% Tween-20) for blocking, membrane was incubated at 4 °C overnight with primary antibodies; anti-Flag (1:1000, Sigma, Cat#F1804), anti-GFP (1:2000, Abcam, Cat#ab290), anti-TDP-43 (1:1000, Proteintech, Cat#12892-1-AP), anti-Lamin B (1:1000, Abcam, Cat#ab16048), anti-GAPDH (1:1000, Santa-Cruz, Cat#sc-32233), and anti-c-Abl (1:1000, CST, Cat#2862) antibodies at 4 °C overnight followed by incubation with HRP-conjugated rabbit of mouse secondary antibodies (1:50,000; GE Healthcare) and HRP-conjugated mouse of donkey secondary antibodies (1:10,000; GE Healthcare) for 1 h at room temperature (RT). Immunoblot signals were visualized by enhanced chemiluminescence (Thermo Scientific, Rockford, IL, USA). The membranes were reprobed with HRP-conjugated anti-β-actin antibody (1:40,000, Sigma, Cat#A3854).