Differential in vivo Thiol Trapping with N-ethylmaleimide (NEM) and 4-acetamido-4'-maleimidylstilbene-2,2'-disulfonic acid (AMS)

Materials and Reagents

1. Bacteria (this protocol works for E. coli MG1655 and Vibrio cholerae O395 strains, I didn’t try other bacteria)
   
2. HOCl (Sigma-Aldrich)
   
3. Trichloroacetic acid (TCA)
   
4. Urea
   
5. Tris-HCl
   
6. EDTA
   
7. SDS
   
8. N-ethylmaleimide (NEM)
   
9. DTT
   
10. 4-acetamido-4'-maleimidylstilbene-2,2'-disulfonic acid (AMS)
    
11. Polyclonal antibodies against your protein and appropriate secondary antibodies
12. LB media (see Recipes)
    
13. DAB buffer (see Recipes)

Equipment

1. Centrifuges
   
2. Western blotting equipment
   
3. Spectrometer
   

Procedure