Ribophorin lysis in SDS

Lysis buffer (50 ml)

2% SDS (5 ml 20%)

25 mM Tris pH:7.4 (1,25 ml 1M)

100 mM NaCl (1 ml 5 M)

3 mM EDTA (300 mcl 0.5M)

Protease Inhibitor Cocktail

Dilution buffer:

0.2% SDS (2,5 ml 20%)

25 mM Tris pH:7.4 (6,25 ml 1M)

100 mM NaCl (5 ml 5 M)

3 mM EDTA (1,5 ml 0.5M)

1,25 % Triton X100 (3,125 ml)

Lysis of cell in 300 microl, boil 3 min, sonicate (3 shots max), dilute with 1 ml dilution buffer. After immunoprecipitation wash beads with dilution buffer 3 times and 1 time

Tris 10mM pH:7.4 (H₂O lavaggi)