DEAE Anion Exchange Protein Purification Protocol

Running DEAE

Solutions needed
~ 100mL 40mm Tris, pH 7.8

~ 60ml 40mm Tris, pH 7.8 + 0.6 m NaCl

DEAE Anion Exchange Protein Purification Protocol

Depending on how powerful your protein bound, you may want to use a higher molarity of NaCl for your second solution

sample Dialyze
Stir in cold room
Change outside the buffer before the leave (supposed to be stirred at least one hour)
Change outside the buffer again the next day and stir at least one hour
Spinning down dialyzed sample to make sure theres no pellets
If there is a pellet, resuspend in a high-salt solution and run some gel
Pour the rest into the column

Prepare column
Pour 8 ml beads to the tube
Wash 3x with water, 1x with Tris alone, 1x with high salt Tris +
Put enough of Tris-only solution to resuspend beads again (~ 16mL if 8 mL of beads)
pour it into the column and let it run
Pour some of Tris-only solution and let it run a little column in the same tube that ran into the sample
Running multiple samples on the gel
Run Tris-only solution through the column til OD280 decreased to ~ 0.02 to 0.01.
Start the gradient (high-salt solution on the left, low-salt solution to the right if the apparatus gradient starting from the right side; on the contrary if the fluid flow from the first left)
–push to get rid of air bubbles
Keeping the room on the right stirring
Columns must be emptied at the same rate that the solution enters

Read OD280 of each of 40 tubes
Pour back into the tube right after each reading