Hi Liew,
There have already been some very good suggestions. I agree with Tim
Gruene that a great starting point is to test the diffraction properties
of your crystal at room temperature. This can serve as a baseline for
comparing/evaluating cryoprotecting agents and methods.
You can also test your potential cryoprotection solutions to see if
they freeze clear or even take a few X-ray snapshots of them to confirm
there are no ice rings.
There are lots of publications that can be helpful. One very helpful
reference is:
Garman, E.F. and Doublie, S.
Cryocooling of Macromolecular Crystals: Optimisation Methods.
Methods in Enzymology (2003) 368, 188-216.
Be aware that, as mentioned previously, the method for freezing can
make a difference (e.g. freezing in cold-stream vs. plunging in
nitrogen). An obvious difference is different cooling rates (you can
find references for this) or less obvious reasons, for example
differences in dehydration that occur during longer/shorter transfer
through air from drop to cold-source for either method.
Finally, you can check out the database for cryoprotecting solutions:
http://idb.exst.jaxa.jp/db_data/protein/search-e.php
Good luck,
-Andy
--
=============================================
Andrew T. Torelli Ph.D.
Postdoctoral Associate
Laboratory of Steven E. Ealick
Department of Chemistry and Chemical Biology
Cornell University
=============================================
On 4/24/2009 10:44 AM, Liew Chong Wai wrote:
Hi all
Thanks for your precious suggestions and ideas.
The crystallization buffer condition is 0.1M BIS-TRIS pH 5.5, 0.2M
MgCl.6H2O, 35% PEG3350
Now, my crystal seem ok in 20% ethylene glycol, but only after a couple
minutes of dehydration at room temperature. For sure, i will try other
cryoprotectant that was suggested here.
I just wondering why MPD kills the crystal.
Many thanks
*LIEW*
--
=============================================
Andrew T. Torelli Ph.D.
Postdoctoral Associate
Laboratory of Steven E. Ealick
Department of Chemistry and Chemical Biology
Cornell University
=============================================
