Glutathione Beads Regeneration

1. Wash glutothione beads with 3 CV of high pH buffer (0.1 M Tris-HCl + 0.5 M NaCl, pH 8.5).
2. Wash beads with 3 CV of water
3. Wash beads with 3 CV of low pH buffer  (0.1 M sodium acetate + 0.5 M NaCl, pH 4.5).
4. Repeat from step 1 and step 3 three times.
5. Equilibrate with 3 CV of 1X PBS.
6. To regenerate really dirty beads, wash beads with 2 X 6 M guanidine hydrochloride.
7. Immediately wash with 5 bed volumes of 1X PBS.
8. To remove hydrophobically bound substances, wash beads with 3–4 bed volumes of 70% ethanol or with 2 bed volumes of a non-ionic detergent (conc. 0.1%), immediately followed by a wash with
5 bed volumes of 1X PBS.
9. Wash with 3 CV of 20% ethanol.
10. Store at +4 °C.
11. Equilibrate the beads with 1X PBS before reuse.