Materials and Reagents

1. EDTA free protease inhibitors (F. Hoffmann-La Roche, catalog number: 05892988001 )
   
2. Proteasome inhibitor MG-132 (Calbiochem^®, catalog number: 474790-5MG )
   
3. Phosphate buffered saline (PBS)
   
4. Tris
   
5. Ni-NTA-Agarose beads (QIAGEN, catalog number: 30210 )
   
6. Urea
   
7. Guanidinium-HCl ( Sigma-Aldrich, catalog number: G4505-25G )
   
8. Imidazole (Sigma-Aldrich, catalog number: 12399 )
   
9. Triton X-100
   
10. Glycerol
    
11. β-mercaptoethanol
    
12. SDS
    
13. Bromophenol blue
    
14. Lysis buffer (see Recipes)
    
15. Wash buffer (see Recipes)
    
16. Elution buffer (see Recipes)
    
17. 4x laemmeli buffer (see Recipes)
    

Equipment

1. Centrifuges
   
2. Sonicator
   
3. Western blotting equipment
   
4. Tissue culture plates
   
5. 1.5 ml eppendorf tube
   
6. 15 ml falcon tube

Procedure