Anti rabbit igg fc

Total proteins of cells were extracted using RIPA lysis buffer (Beyotime) with Protease Inhibitor (Roche) and Phosphatase Inhibitor (Roche). Protein samples were separated in sodium dodecyl sulfate (SDS)-PAGE and transferred to polyvinylidene fluoride (PVDF) filter membranes (Millipore, USA) for immune-hybridization. After 1 h of blocking in PBST (phosphate buffer saline containing 0.05% Tween-20 and 5% non-fat milk powder), the membranes were incubated with one of the following primary antibodies with corresponding concentration: RNF8 (Santa Cruz Biotech, 1:500), YBX1 (Cell Signaling Technology, 1:1000), HA (Convance, 1:1000), Secondary antibodies were Horseradish peroxidase (HRP)-conjugated anti-mouse IgG (ZB-2305, ZSGB-Bio, 1:4000) or anti-Rabbit IgG(Fc) (ZB-2301, ZSGB-Bio, 1:4000). Subsequently, band visualization was performed by electro-chemiluminescence (ECL) and detected by Digit imaging system (Thermo, Japan), the gray level of the bands was quantitated by ImageJ software.

Total proteins were extracted with RIPA lysis buffer (Beyotime) with Protease Inhibitor Cocktail (Roche) and Phosphatase Inhibitor Cocktail (Roche). Protein samples were separated in sodium dodecyl sulfate (SDS)-PAGE and transferred to polyvinylidene fluoride (PVDF) filter membranes (Millipore, USA). After blocking in phosphate buffered saline (PBS) containing 0.05% Tween-20 and 5% non-fat milk powder, the membranes were incubated with the following primary antibodies: RNF8 (Santa Cruz Biotech, 1:500), EMT kit (Cell Signaling Technology, 1:2000), beta-actin (Santa Cruz Biotech, 1:4000), Secondary antibodies were Horseradish peroxidase (HRP)-conjugated anti-mouse IgG (ZB-2305, ZSGB-Bio, 1:4000) or anti-Rabbit IgG(Fc) (ZB-2301, ZSGB-Bio, 1:4000). Subsequent visualization was detected by Digit imaging system (Thermo, Japan), the gray level of the bands was quantitated by ImageJ software.