[Protein-analysis] Re: preparing proteins for lyphilization
(by sumida from bbri.org)
Fri Jul 11 21:06:57 EST 2008
Thank you for your response to my post. I have found this to be
generally true, and I remember that it was told to me sometime ago,
but there are some questions I wonder about.
One would think that the pH could drop significantly during this
process; I normally use a 5 to 10mM buffer concentration which would
not have a significant buffering capacity. I suppose the reasoning is
that HCO3 would eventually convert to CO2 and be lost. But what
happens to the solution freezing point and consequently the vapor
pressure. Wouldn't it become more and more difficult to sublimate the
dissolved gases as the solute concentration increases?
I cannot seem to find any type of literature reference for this piece
of information. Do you know if anyone has performed any physical
characterization of this process? I have been searching pubmed and
JSTOR but have yet to find anything.
On Jul 7, 11:59 am, d... from no.email.thankstospam.net (DK) wrote:
> In article <279a3698-1bd7-4ec3-8887-62e9efc4f... from t12g2000prg.googlegroups.com>, jps <sum... from bbri.org> wrote:
> >I think it is common practice to dialyze proteins in an ammonium
> >bicarbonate solution at pH=8. Presumably this is a way to maintain
> >the protein with some salt to stabilize it while in solution, but not
> >have any salt present after this protein solution is lyophilized since
> >the ammonia sublimates and carbon dioxide is released during freeze
> >drying step.
> >However, I am wondering if this is actually accurate. Can anybody
> >comment on this procedure?
> This is actually accurate. Of course, some proteins survive lyo just
> fine while others tend to die. You never know until you try. Addding
> some sugars into protein solution to be lyophilized frequently helps
> because they act as cryoprotectants. Trehalose is very popular but
> sucrose is always worth trying (trehalose is usually less pure).
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