Re: [ccp4bb] cryoprotectant for crystals in isopropanol
Not that all proteins are created alike, but we had a protein crystallize in a similar situation (PEG/20%IPA). I don't remember the exact details, but we did have good luck using just glycerol as a cryoprotectant. That is, I made up the well buffer and added to that well buffer 20-25% glycerol (in addition to IPA/PEG), and moved the crystal into that. We had no issues with cracking or anything, and data collection went well. The first thing I tried worked, so I don't have a lot of trial information for you. You could probably reduce the glycerol a bit with no worries, but I don't know. Good luck Dave shivesh kumar wrote: Dear all, The question is regarding the cryoprotectant.We have crystallized a protein in 20% PEG 500 with 10% isopropanol.What should be the cryo we should try for data collection.Also,what percentage of PEG 500 is sufficient enough as a cryo for data collection. We are also trying to crystallize it with higher PEG's also like 1K, 2K, 4K.Lets C.Since the condition is having isopropanol also and it is reported that mounting with isoprop is very difficult and it evaporates as soon as we open the slides. Thanx in advance... Shivesh Kumar
Re: [ccp4bb] cryoprotectant for crystals in isopropanol
Another anecdote for you: I had crystals that grew in 15% PEG 2000 and 15% isopropanol. Cryos with glycerol melted the crystals. The best cryo I found was with 15% PEG 2000, 15% isopropanol, and 20% PEG 400. It's hard to predict how your crystals will behave with different cryoprotectants-- hopefully you have enough crystals to try several different conditions. Evette S. Radisky, Ph.D. Assistant Professor and Associate Consultant II Mayo Clinic Cancer Center Griffin Cancer Research Building, Rm 310 4500 San Pablo Road Jacksonville, FL 32224 (904) 953-6372 (office) (904) 953-0046 (lab) From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of shivesh kumar Sent: Wednesday, March 19, 2008 3:34 AM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] cryoprotectant for crystals in isopropanol Dear all, The question is regarding the cryoprotectant.We have crystallized a protein in 20% PEG 500 with 10% isopropanol.What should be the cryo we should try for data collection.Also,what percentage of PEG 500 is sufficient enough as a cryo for data collection. We are also trying to crystallize it with higher PEG's also like 1K, 2K, 4K.Lets C.Since the condition is having isopropanol also and it is reported that mounting with isoprop is very difficult and it evaporates as soon as we open the slides. Thanx in advance... Shivesh Kumar
