Flag m2 monoclonal antibody f1804

Whole cell protein extracts were resolved on SDS-PAGE and proteins were transferred to nitrocellulose membranes. After blocking for 1 hour at room temperature in 5% milk in PBS/0.1% Tween-20, membranes were incubated overnight at 4oC with the indicated primary antibodies AR (441), AR (N-20), Ubiquitin (P4D1 and FL76), 1:1000 dilution, Santa Cruz Biotechnology, Santa Cruz, CA. AR-V7 (AG10008, Mouse monoclonal antibody, 1:1000 dilution, precision antibody); FLAG® M2 monoclonal antibody (F1804, 1:1000 dilution, Sigma-Aldrich, St. Louis, MO); AKR1C3 (A6229, 1:1000 dilution, Sigma-Aldrich, St. Louis, MO); AKR1C3 (11194–1-AP, 1:1000 dilution, Proteintech Group, Inc); Tubulin (T5168, Monoclonal Anti-α-Tubulin antibody, 1:5000 dilution, Sigma-Aldrich, St. Louis, MO). Tubulin was used as loading control. Following secondary antibody incubation, immunoreactive proteins were visualized with an enhanced chemiluminescence detection system (Millipore, Billerica, MA).

Whole cell protein extracts were incubated for 16 hours at 4°C with the indicated primary antibodies AR (441), AR (N-20), AR (C-19), HSP70 (F-3 and H-300), STUB1(H231 and G-2), HA (F-3), Ubiquitin (P4D1 and FL76), 1:1000 dilution, Santa Cruz Biotechnology, Santa Cruz, CA; STUB1 (C3B6, 1:100 for IP, Cell Signaling antibody); AR-V7 (AG10008, Mouse monoclonal antibody, 1:1000 dilution, precision antibody); FLAG® M2 monoclonal antibody (F1804, 1:1000 dilution for western blot, 1:200 for IP, Sigma-Aldrich, St. Louis, MO); Tubulin (T5168, Monoclonal Anti-α-Tubulin antibody, 1:5000 dilution, Sigma-Aldrich, St. Louis, MO). Following secondary antibody incubation, immunoreactive proteins were visualized with an enhanced chemiluminescence detection system (Millipore, Billerica, MA). The bands were quantified by ImageJ.