Dialysis device

BL lo1 at ntlworld.com
Thu Nov 15 11:09:09 EST 2001


>> 
>> This erroneous point of view appears to be widely held and is known to
>> be a cause of many mistakes. I've seen a grad student who was
>> seriously attempting to separate 50 kDa protein from 20 KDa contaminant
>> this way (a completely hopeless and futile exercize). Truth is, cut-off
>> value in both dialysis and ultrafiltration refers to a _retention_. As such,
>> a cut off of 30 kDa does not mean that a 10 kDa molecule with go
>> through with an efficiencty much higher that that of 30 kDa. All
>> such a value says is that >95% of 30 kDa molecule will be
>> retained (and less of 10 kDa molecule will be retained). It is prudent
>> to keep in mind, for example, that even salts and things like glycerol
>> and sucrose are concentrated to some extent by 10 kDa cut-off
>> Centricons.
>> 
>> DK
> 
> Point taken DK, 
> I was simplifying for the sake of message length.  Also, I believe that
> microcons come with instructions about cutoffs, how to do buffer
> exchanges, etc.  The original message did not say what was to be removed
> by dialysis and I assumed that BL already understood the theory of
> dialysis. I guess I should have said, "most of things larger then the
> mol wt cut off will be retained, and almost of the salts will pass
> through".
> 
> While there will be 'some' retention of salts, if you dilute say a 50 ul
> sample into 500 ul, bring it down to 50 ul and then repeat, would not
> this work as well as dialyzing against several buffer changes?
> Except for removal of amino acids from large proteins, or free dNTPs
> from probes, I would not consider dialysis or centricons a good choice
> for seperating macromolecules from each other.

Thank you for all your suggestions. I require the filter for a protein
refolding experiment, so in essence it's for the gradual removal of urea (or
other denaturants like GdHCl). I found this DispoDialyzer seems to be
suitable. I am getting a sample to have a look.

BL




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