Re: [ccp4bb] Questions about cryoprotectant

[Roger Rowlett]

You have options: 1. Paratone-N (rarely works for me when other methods don't also work) 2. ML + 25-30% glucose (one of my favorite and reliable methods) 3. ML + 25-30% EG, maybe DMSO PEG400 is not likely to work here because its solubility in high sulfate solutions is limited. In 2 M AmSO4, you can only dissolve about 4% PEG-400. Typically flash-cooling increases mosaicity to some degree. If you have crystals to burn, you can try flash-annealing them in the cryostream by interrupting the flow with a credit card for about a 5-count. For some crystals this is magic in reducing mosaicity of flash-cooled crystals, but it *never* works for me :( Cheers. ycheng wrote: Hi, I am trying to find an appropriate cryo-condition for my protein crystals. The mother liquid is 2-2.5M Ammonium phosphate dibasic 100mM TrisHCL pH8. The room-temperature diffraction looks not bad (mosaicity 0.8, resolution 2.6) But the diffraction turned to be very mosaic if I freeze the crytals in the absence of cryos or in the presence of mother liquid plus different concentration of glycerol (5%,10%,15%.20%). I don't think the ice formation is the problem since I didn't see any ice by my eyes or ice diffraction in the presence or absence of cryos. Also, I didn't see any cracks on my crystals when I transfered them to the cryo conditions I have already tried. My question here is: 1)what's the role of cryo? I know it helps prevent ice formation. Based on my case, it looks like cryo might also help to keep the crytal packing good when frozed. 2) What do I need to do to find a good cryo? What in my mind is to try other cryos like sucrose, PEG400, ethylene glycol. Thanks a lot for your attention! Yuan -- Roger S. Rowlett 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