CRYO PROTECTANT

Ana Gonzalez gonzalez at esrf.fr
Wed Nov 8 13:14:16 EST 1995


Hakon Hope <hhope at ucdavis.edu> wrote:
>
>    Response to  Ana Gonzalez (gonzalez at esrf.fr)
>
>If your crystal was suspended in a thin solven film,
>evaporation of water could be fast enough to change the
>composition so that effectively the film was cryoprotected.
>
This could certainly be a possibility, although it takes about the same
time to fish out the crystal from the mother liquour and put it in the 
cold air stream as to plunge it into liquid N2 (both were done as quickly 
as possible). However I agree, in a proper experiment this should be monitored.

BTW, if someone wants to have a go at trying it, the lyso crystals were 
tetragonal grown in  acetate pH 4.7 -I think! using NaCl as precipitant.
I can dig out the detailed recipe if someone needs it...Note there
is no PEG in the mother liquour as  in other "self-protecting" crystals...

>The normal behavior would be for ice to form from the mother
>liquor, both in liq N2, liq propane, or in a cold stream.

Yes, but from the diffraction pattern quality point of view it makes a
difference if it is vitreous (hyperquenched)  ice, or crystalline ice. 
>
>Do you know that the ice formed on cooling in liq N2,
>and not during the transfer to the diffractometer?
>
I was using a transferring device which  keeps the crystal in liquid nitrogen
until it is centered in the cold air stream. It was designed at the EMBL
Heidelberg and based on the "extended goniometer arc" idea. It was 
possible to take the crystals off the air stream into liquid nitrogen
and then remount them again and you don't get any (crystalline ;) icing.

I still think a little hesitation holding the crystal above the liquid N2
(while freezing it directly in the liquid) could explain the crystal ice
formation.

Another thing that differs from your experiment is that you used a really
thick glue coat over your probe (to simulate the cryoprotectant 
shielding effect I guess) while the mother liquour coat is extremely thin.
Could it just be that the cold air stream cools down the surface (ie
the mother liquour) faster than the inmersion, even if the "bulk" lags
behind and cools down slower by the former method?

I think it would be great if it was possible to freeze crystals more often
without cryoprotectant: less background, easier manipulation of the crystals...
However mounting them directly on the cryocooler stream is pretty hairy, it
would be so much easier to pre-freeze them in a cold liquid...
So it will be great news if someone proves I was doing something wrong ;)

Ana
-- 
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     /    \  /  \    /  \      Ana Gonzalez (gonzalez at esrf.fr)
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