User:Tamra L. Fisher/Notebook/Experimental Biological Chem/2012/03/21

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FPLC of MBP-intein.


  1. Inject about 20mL of the dialyzed supernatent onto the anion-exchange column. (except for the first time when only 5mL was injected)
  2. Run 25mL of 50mM Tris (pH~9) over the column.
  3. Start fraction collection. Run a gradient of 50mM Tris from 0M NaCl to 0.5M NaCl over 20 minutes. Collect in 5mL fractions.
  4. Run 25mL of 50mM Tris + 1M NaCl (pH~9.15) over column.
  5. Run 25mL of 50mM Tris (pH~9) over the column.
  6. Repeat until all of the supernatent has been injected and eluted off of the column.


This is the output from the HPLC:

032212 mbp intein purification.JPG