Hello,
I have a question for old school scientists. I read in an old book about protein purification about the vacuum dialysis method for volume reduction and buffer change. When a sample is placed in a dialysis bag and placed into the mouth of a buffer-filled vacuum flask and incubated in mild vacuum.
Out of curiosity, I assembled the installation according to the scheme in Current protocols in molecular biology book, but no matter how I tried, the dialysis bag burst, either when using a vacuum pump, or when using a water jet pump. What could be the reason - were dialysis tubes made better before? Or should the vacuum be very very weak? I would be grateful if you would share your experience, I am very interested in old techniques in protein purification.
Another trick was to wrap the dialysis tubing with dry polyethylene glycol or dry Sephadex, which would suck out the water by osmosis from the inside of the tubing.
Dear Pierre, thank you for your reply!
I have tried that approach too, it works fine although not so quick. I was just surprised that the vacuum dialysis method is often mentioned in old books, say the 70s-80s, as something common. But it was not possible to repeat the method, most likely the point is that the vacuum was too strong.
I never heard of this method. I think it would be very important to make sure there is no air bubble inside the dialysis tubing, because it will expand under vacuum and create enough pressure that the bag might burst.
Just like Adam, I have never heard of this method. In theory, reducing the pressure on the outside of a semipermeable membrane would add to the osmotic pressure gradient, and accelerate osmotic flow out of the tube. The limit, as you found out, is the stability of the membrane. I'd rather trust my precious samples to an Amicon cell instead.
If the main point is to concentrate the solute then just hanging the dialysis bag filled with solution in an air flow like a fume cupboard door will evaporate the solvent and concentrate the solute while the evaporation does keep the sample a bit cool but there are many better methods.
Like @pierre Beguin says sephadex works well but if you dont mind losing some protein then it is much quicker to swell sephadex which excludes your Mol Wt protein directly in the solution when speed matters more than cost
Dear all! Thank you for your reply!
I know that now this method is too archaic and there are many other better methods of concentration and I use them (rarely Sephadex and dialysis tubing, more often amicones and TFF). I'm just interested in old-school methods more from a historical point of view and sometimes I want to reproduce them. Anyway thanks for your comments!
Years ago, our lab had a fast, convenient device to concentrate samples by vacuum dialysis. I think it was called Micro-ProDiCon from Spectrum. As I recall, it required special thick dialysis membranes that could withstand the pressure.
Dear Don, thank you.
Maybe this was the trick, in the thick dialysis tube.
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