TSBD TRYPTICASE SOY BROTH DIALYSATE

Use for producing stable exoenzyme S.

  1. 1 litre
  2. 30 g TSB
  3. 90 ml dH2O
  4. 10 g Chelex 100-400 (other Chelex is ok)
  5. Stir 6 hours at 25° C.
  6. Spin 16,000 g for 30 minutes.
  7. Filter through 0.45 um filter.
  8. Concentrate to almost dry over a PM-30 (PM-10 is ok) Amicon membrane.
  9. Discard concentrate on the filter, the rest is 10X and filtered.
  10. Add nitrilotriacetic acid to 10 mM.
  11. Adjust to pH 7.0.
  12. Autoclave in 2 litre flask. Cool.
  13. Add 20 ml sterile 50% glycerol.
  14. Add 50 ml sterile 2 M MSG (or glutamate, Na salt)
  15. Add sterile MgSO4 to 1 mM