Dialysis of Purified Protein

 

 

  1. WEAR GLOVES ANY TIME YOU ARE DOING A DIALYSIS
  2. Get a large 2000ml beaker or larger if necessary
  3. Estimate the volume of your sample and make a 100 fold excess amount of 10X buffer (If your sample volume is 10ml then make 1000ml of 10X buffer)
  4. Place the dialysis buffer in the large beaker, cover with saran wrap, and place in the cold room until further use.
  5. Get 2 orange dialysis clamps
  6. Get enough dialysis bag to hold all of your sample.  Each millimeter can hold about 1ml so measure out a length equal to the volume of your sample.
  7. Get a new clean razor blade and make one clean cut at the end of the length you re going to use. 
  8. Rinse the orange clamps and the dialysis bag excessively with dH2O.  Rinse the dialysis bag very well so that it becomes hydrated.
  9. Once you have hydrated the dialysis bag you can roll your fingers along one of the ends until it opens.
  10. Once you have one end of the dialysis bag open run lots of dH2O through it to rinse the inside and fully hydrate it.
  11. Take one of the orange clamps and clamp one of the ends so that it is closed shut. 
  12. Fill the entire dialysis bag with water so that you can look for leaks.  Apply a little pressure to the bag to look for small holes and to make sure that the clamp is working properly.
  13. Grab the test tube samples from the cold room that you have decided to use according to your gel.
  14. Use a powered pipette to collect all the fractions that you plan to dialyze and put the entire sample into the dialysis bag.
  15. Clamp the other side of the dialysis bad with the other orange clamp leaving a little room at the end so that the bag is easy to grab and work with.
  16. Dialyze for three changes of buffer each going over 8 hours.