Centrifugal Protein Concentration and Buffer Exchange
Centrifugal Protein Concentration/Buffer Exchange
- Obtain a centrifugal filter (Amicon or Centricon) with a molecular weight cutoff at least half of your protein’s weight.
- If the filter is new, run ~5 mL of nanopure water through before adding your protein mixture.
- Make sure to balance the centrifuge with another centrifugal filter. I typically use an old one. Regular falcon tubes will unbalance the centrifuge even though they are the same weight.
- Add your protein mixture up to the recommended line.
- Spin at 4,000 rpm for ~40 minutes.
- Pour out the filtrate. The supernatant contains your protein.
- Add the rest of your protein mixture or add in the new buffer.
- Continue to add and spin down the new buffer until it is sufficiently exchanged and the protein is concentrated.
- ~50 mL works well
- Be sure not to over concentrate and aggregate your protein.
- Keep the filter wet with 20% ethanol when it is stored.