Dear all,
Does anyone have experience in comparing normal dialysis x ultrafiltration in terms of protein loss?
I am extracting membrane proteins from Leishmania donovani and I have to concentrate my samples upon completion of the experiment. I have been using ultrafiltration devices of 3 kDa MWCO (vivaspin 6 ml combined with amicon ultra 0.5 ml) to concentrate my samples as well as to get rid of the detergent (octyl glucoside). They work fine, I do get more concentrated samples, despite some protein loss.
Therefore I am now wondering whether, instead of ultrafiltration (that concentrates my samples as well as eliminate salts and the detergent) dialysing my sample (to get rid of the detergent) and then using another method to precipitate/ concentrate proteins like ethanol precipitation would be a more effective one.
Thanks and best regards
Bruno Hinckel
Hello Bruno,
For protein concentration ultrafiltration works well. You may also use precipitation techniques with various chemicals. No matter what method you use you will certainly have a loss, there is no perfect system that will give you 100% recovery.
As far as dialysis is concerned it is often used to rid protein solution of unwanted chemical components that may interfere with downstream experiments. It does not concentrate proteins. In contrast it often dilutes proteins due to increased volume of resulted solution. Afterwards the solution has to be concentrated.
Good luck!
Chao
Hello Chao,
thank you very much for your answer.
You are right, I have formulated my question not the best way. To be more precise, I am wondering whether the protein loss using dialysis would be less significant than the protein loss using ultrafiltration (due to higher adherence of proteins to the filter membrane due to the high centrifugal forces). As you said, there will be a loss in any system, but I am now wondering whether the loss (and not protein concentration!) of protein using dialysis would be smaller than that of using ultrafiltration.
If this shall be the case, I might use dialysis to get rid of my detergent and the try ethanol precipitation to concentrate the proteins.
Best regards
I have not done a well controlled experiment on those two methods for protein recovery, so I can not answer your question. I am sure somebody has done it, and I hope you will get an answer soon!
Chao
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