Hi! I need to centrifuge using amicon ultra-15 to concentrate my protein. I have some doubts. First of all, if I can do it at 4 ºC, if it would be better for proteins, although in the protocol it is said to centrifuge at room temperature.
In addition we only have got a centrifuge where the amicon enter without cap, do you think this would be a problem and it would be better to find another centrifuge? I have thought to centrifuge with parafilm...
And finally about re-use...the best opcion would be after 1st use wast with water, then with 70 % ethanol and finally store at 4 ºC with water? Or do I include a centrifugation with water or 70 % ethanol?
Thank you!
hi
1- Before starting you should wash the amicon with DDW and centrifuge
.
2- According to the protocol of the company listed in the centrifuge.
3- you can do it at 4c but The time for centrifuges will be longer 1-2 hour (Filter pores become smaller At low temperature)
4- finally wash the filter by DDW + centrifuge and save the filter in 20% ethanol .
good luck
Dear Damaris,
no problem to centrifuge at 4 °C and without cap (we have done it occasionally by ourselves).
Best,
Christoph
Damaris-
1) yes, filtration can be performed at 4C (note that the filtration time would be longer versus spinning at room temperature).
2) You could certainly place Parafilm on top of device during centrifugation, but do not seal too tightly or flow may not initiate (cap w/device is designed to allow air into device during spin).
3) Amicon Ultra's are single use only, however, some researchers have had success by spin/rinse/washing 0.1 N NaOH followed by a second spin of buffer or Milli-Q® water, but CAUTION: Do not allow the membrane in Amicon® Ultra filter devices to dry out once wet. Ethanol would be used secondary if sanitization desired by applying 70% ethanol to filter component, incubating ~30 minutes, spinning ~50% of volume of ethanol through device (discard remaining retentate and filtrate), and follow up with sterile Milli-Q® water rinse (method as noted previously) and final filling with Milli-Q® water to keep hydrated until next use.
Please reach out if you need more information
Janet Smith
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