Re: [ccp4bb] Cryo solution for crystals grown in magnesium formate
You can increase the formate concentration and it will be a cryoprotectant on its own. Test increasing amounts until you find the concentrationthat freezed clear as glass and the transfer your crystal to this. We did this with sodium formate at 7 M quite easily. On Tue, Dec 17, 2013 at 2:59 AM, Tim Gruene wrote: > -BEGIN PGP SIGNED MESSAGE- > Hash: SHA1 > > Dear Junyu, > > You can test cryo conditions conveniently in the absence of your > crystals if you have access to e.g. an inhouse machine. Prepare your > solutions with the usual suspects (PEG400; PEG400+glycerol; > butanediol; Na Malonate etc. pp.) at various concentrations, pick up a > large as possible drop with a nylon loop and take a shot to see if you > have ice rings. > Try to be on the safe side with respect to the decision about the > minimal concentration because the presence of the crystal might give a > slightly different result. > > Best, > Tim > > On 12/16/2013 10:36 PM, Xiao, Junyu wrote: > > Dear all, sorry if this topic does not interest you. I wonder > > whether anyone has experience with freezing crystals grown in ~0.2 > > M Magnesium Formate. Garman and Mitchell suggested that "A major > > anomaly is solution 44, 0.2 M magnesium formate, which requires 50% > > glycerol for cryoprotection" in their 1996 paper (J Appl. Cryst. > > 29, 584-587). Since 50% glycerol is kind of harsh, I wonder > > whether anyone has tried alternative cryo protectant. Your kind > > help will be highly appreciated. > > > > Best regards, Junyu > > > > --- Junyu > > Xiao, Ph.D. University of California, San Diego Leichtag Room 283 > > 9500 Gilman Drive, 0721 La Jolla, CA 92093-0721 Lab phone: > > 858-822-0684 > > > > > > > > - -- > - -- > Dr Tim Gruene > Institut fuer anorganische Chemie > Tammannstr. 4 > D-37077 Goettingen > > GPG Key ID = A46BEE1A > > -BEGIN PGP SIGNATURE- > Version: GnuPG v1.4.12 (GNU/Linux) > Comment: Using GnuPG with Icedove - http://www.enigmail.net/ > > iD8DBQFSsBJ/UxlJ7aRr7hoRApCiAJ0b3LyawgscAsIzwFUUHNDqRSplPwCeOZPv > pF9iloWrCgMjIwo4hKlAgN8= > =l4tU > -END PGP SIGNATURE- >
Re: [ccp4bb] Cryo solution for crystals grown in magnesium formate
I find that odd because the first structure I ever worked on was a DNA oligonucleotide grown in 0.2 M magnesium formate and it did not require cryoprotection at all when flash cooled directly in the cryostream. I learned from Ned Seeman's group a long while back that Mg2+ itself worked reasonably well as a cryoprotectant at high concentrations. We routinely freeze DNA crystals with 120 mM Mg formate and 10% MPD. --paul On 12/16/2013 04:36 PM, Xiao, Junyu wrote: Dear all, sorry if this topic does not interest you. I wonder whether anyone has experience with freezing crystals grown in ~0.2 M Magnesium Formate. Garman and Mitchell suggested that "A major anomaly is solution 44, 0.2 M magnesium formate, which requires 50% glycerol for cryoprotection" in their 1996 paper (J Appl. Cryst. 29, 584-587). Since 50% glycerol is kind of harsh, I wonder whether anyone has tried alternative cryo protectant. Your kind help will be highly appreciated. Best regards, Junyu --- Junyu Xiao, Ph.D. University of California, San Diego Leichtag Room 283 9500 Gilman Drive, 0721 La Jolla, CA 92093-0721 Lab phone: 858-822-0684
Re: [ccp4bb] Cryo solution for crystals grown in magnesium formate
-BEGIN PGP SIGNED MESSAGE- Hash: SHA1 Dear Junyu, You can test cryo conditions conveniently in the absence of your crystals if you have access to e.g. an inhouse machine. Prepare your solutions with the usual suspects (PEG400; PEG400+glycerol; butanediol; Na Malonate etc. pp.) at various concentrations, pick up a large as possible drop with a nylon loop and take a shot to see if you have ice rings. Try to be on the safe side with respect to the decision about the minimal concentration because the presence of the crystal might give a slightly different result. Best, Tim On 12/16/2013 10:36 PM, Xiao, Junyu wrote: > Dear all, sorry if this topic does not interest you. I wonder > whether anyone has experience with freezing crystals grown in ~0.2 > M Magnesium Formate. Garman and Mitchell suggested that "A major > anomaly is solution 44, 0.2 M magnesium formate, which requires 50% > glycerol for cryoprotection" in their 1996 paper (J Appl. Cryst. > 29, 584-587). Since 50% glycerol is kind of harsh, I wonder > whether anyone has tried alternative cryo protectant. Your kind > help will be highly appreciated. > > Best regards, Junyu > > --- Junyu > Xiao, Ph.D. University of California, San Diego Leichtag Room 283 > 9500 Gilman Drive, 0721 La Jolla, CA 92093-0721 Lab phone: > 858-822-0684 > > > - -- - -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A -BEGIN PGP SIGNATURE- Version: GnuPG v1.4.12 (GNU/Linux) Comment: Using GnuPG with Icedove - http://www.enigmail.net/ iD8DBQFSsBJ/UxlJ7aRr7hoRApCiAJ0b3LyawgscAsIzwFUUHNDqRSplPwCeOZPv pF9iloWrCgMjIwo4hKlAgN8= =l4tU -END PGP SIGNATURE-
Re: [ccp4bb] Cryo solution for crystals grown in magnesium formate
On Mon, Dec 16, 2013 at 1:36 PM, Xiao, Junyu wrote: > Dear all, sorry if this topic does not interest you. I wonder whether > anyone has experience with freezing crystals grown in ~0.2 M Magnesium > Formate. Garman and Mitchell suggested that "A major anomaly is solution > 44, 0.2 M magnesium formate, which requires 50% glycerol for > cryoprotection" in their 1996 paper (J Appl. Cryst. 29, 584-587). Since > 50% glycerol is kind of harsh, I wonder whether anyone has tried > alternative cryo protectant. Your kind help will be highly appreciated. > Another good reference: http://journals.iucr.org/j/issues/2002/05/00/do0015/index.html It suggests 35% PEG 400, 30% ethylene glycol, or 30% of whatever "PG" means (based on the rest of the paper I suspect propanediol, but the abbreviation doesn't really make sense - perhaps Eddie Snell can clarify). There are of course many other good cryoprotectants beyond those evaluated in the paper; personally, I'm a big fan of xylitol (which I believe will work in lower concentrations - at least with some conditions), but what really matters is what the crystals can tolerate. Note that these estimates are using very strict criteria - you can often get away with less cryoprotection if you are very good at freezing crystals and/or willing to tolerate some increased background. But I wouldn't try this until you've determined that your crystals can't handle the recommended amounts. -Nat
Re: [ccp4bb] Cryo solution for crystals grown in magnesium formate
Junyu, I haven't tried it personally with this particular solution, but I have found that 30% glucose can pretty much cryoprotect any condition I have tried it with. If necessary, add cryoprotectant solution (mother liquor + 30% glucose) gradually to minimize osmotic shock and potential cracking of crystals. You can try this without crystals to see if the solution vitrifies as a clear solid in liquid nitrogen. If it freezes clear, it is very likely to work fine with your crystals. Cheers, ___ Roger S. Rowlett Gordon & Dorothy Kline Professor Department of Chemistry Colgate University 13 Oak Drive Hamilton, NY 13346 tel: (315)-228-7245 ofc: (315)-228-7395 fax: (315)-228-7935 email: rrowl...@colgate.edu On 12/16/2013 4:36 PM, Xiao, Junyu wrote: Dear all, sorry if this topic does not interest you. I wonder whether anyone has experience with freezing crystals grown in ~0.2 M Magnesium Formate. Garman and Mitchell suggested that "A major anomaly is solution 44, 0.2 M magnesium formate, which requires 50% glycerol for cryoprotection" in their 1996 paper (J Appl. Cryst. 29, 584-587). Since 50% glycerol is kind of harsh, I wonder whether anyone has tried alternative cryo protectant. Your kind help will be highly appreciated. Best regards, Junyu --- Junyu Xiao, Ph.D. University of California, San Diego Leichtag Room 283 9500 Gilman Drive, 0721 La Jolla, CA 92093-0721 Lab phone: 858-822-0684
[ccp4bb] Cryo solution for crystals grown in magnesium formate
Dear all, sorry if this topic does not interest you. I wonder whether anyone has experience with freezing crystals grown in ~0.2 M Magnesium Formate. Garman and Mitchell suggested that "A major anomaly is solution 44, 0.2 M magnesium formate, which requires 50% glycerol for cryoprotection" in their 1996 paper (J Appl. Cryst. 29, 584-587). Since 50% glycerol is kind of harsh, I wonder whether anyone has tried alternative cryo protectant. Your kind help will be highly appreciated. Best regards, Junyu --- Junyu Xiao, Ph.D. University of California, San Diego Leichtag Room 283 9500 Gilman Drive, 0721 La Jolla, CA 92093-0721 Lab phone: 858-822-0684
