Equipment:
- C18 Spec tips, Agilent, cat #A57203
- p200 and p1000 pipetmen
You can use a p1000 pipetman to wash and elute spec tips. For each wash, add the required volume into the top of the spec tip, then press a p1000 tip firmly into the spec tip and depress the plunger to drive the liquid through the device.
- Wet the spec tip by pushing through 200 µl of HPLC grade methanol.
- Wash the spec tip with 3 x 200µl 5% acetonitrile/5% formic acid.
- If your sample does not already contain an acidic ion pairing agent, add formic acid to your sample to 5%. NOTE: Do not pipet concentrated formic acid with plastic pipet tips; use glass.
- Push the sample through the spec tip. You may pass the sample through twice, washing with 5% acetoniltrile/5% formic acid between each pass, if you would like to maximize binding.
- Wash with 3 x 200 µl 5% acetonitrile/ 5% formic acid.
- Elute with 2 x 100 µl 80% acetonitrile/5% formic acid.
- Speed vac the sample to dryness.
Reagents:
- HPLC grade methanol
- 5% HPLC grade acetonitrile/5% formic acid
- 80% HPLC grade acetonitrile/5% formic acid
- Make solutions with MilliQ or better water. NOTE: Do not pipet concentrated formic acid with plastic pipet tips; use glass.
You may also access this protocol via a PDF for easy printing here.