Andrea L Edwards wrote:
Hi all,
What are the most successful methods you know of for dehydrating a crystal
prior to freezing it? I am trying to push the resolution of my crystals.
Thanks,
Andrea
First of all, be aware that not all crystals are improved by
dehydration. Some need to be drier, some need to be wetter.
If you have long or large crystals that extend outside
the loop, you can get a clue by comparing diffraction at the
outside tip (driest) with the center of the loop (wettest).
We've seen dramatic differences in both directions, and
the ones that diffract best outside (beef orthorhombic
cyt bc1) were greatly improved by dehydration, while for
the ones that diffract best in the center (chicken complex II)
we got the best diffraction when they were so wet we started
to get ice rings. Dehydration destroyed them.
Our best dehydration results were with simply holding the
crystal in the air 30 sec to 2 min after mounting before
plunging into LN2. Also something about pushing the crystal
to the edge of the drop where the peg is getting sticky
before fishing. be aware the result may depend on relative
humidity and temperature.
My grand plan for optimizing
dehydration (if the FMS is not available) was to mount
crystals on cryocap pins and screw the caps into vials
containing 0.5 ml of different concentrations of glycerol.
(Pin being short enough that the crystals are suspended
in the air over the solution)
Wait for a few hours at RT or 4* then unscrew the cap and
rapidly freeze. Never really got good results though.
And if the humectant is really wet, the crystal soaks
up so much water it falls out of the loop into the
humectant!
Ed
