I am planning to concentrate phages in wastewater with Amicon Ultra-15.
I will concentrate 40 mL of the sample by repeating centrifuging for four times (10 mL each time)
I have learned that the membrane shouldn't get dried, but i don't understand why. Could anyone tell me the reason?
Also how much should I keep it wet? Should I always keep the membrane soaked in water? Or, can't it contact the air at all?
Thank you Zaizhi Yu for your interest in the Ultracel® membrane in the Amicon® device. Basically, the membrane is composed of layers-the UF skin (that does the work and very thin) and the support material that gives its strength. Once the membrane is wet out, the humectant (i.e. glycerin) that keeps the skin "moist" is washed away. Once the membrane starts to dry out, the thin "skin" layer starts to shrink and separate away from the support making the performance suspect.
You should be able to spin multiple times, but keep the time between spins shorter (not longer than 1-2 hrs if possible, and capped). The membrane just needs to be kept moist in device (Amicon panels covered) for experiments taking longer. Air exposure is not an issue.
Please reach out if you have any more questions.
Janet Smith
Found this article you may be interested in.. Janet Smith
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Zaizhi, my suggestion would be ...
Amicon® Ultra 12 ml/15 mL (fixed angle/swinging bucket). Centrifuge until about 2-3 mL remaining. Add more sample to device to bring to device capacity (12ml/15 mL). Transfer filtrate to a new tube between steps. Mix/pulse on vortex briefly (medium), and spin again until 2-3 mL remaining, etc. until last run when you can bring down the final volume to
Janet Lee Smith
I don't know how to thank you, Janet.
I will carry out your protocol this week and see how much the recovery rate will be improved! I've got excited.
Also, I have one more quick question. Why does 1000xg is better?
I intuitively feel two possivilities:
1. Viral particles (protein) will be distorted by the strong pressure.
2. Viral particles will stick to the membrane by the pressure.
Do my understanding correct?
Yes, gentler on the spin conditions prevents the viral particles from embedding themselves deeper into the membrane, and harder to recover. Some customers may go as low as 500 xg, but usually time is money.
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