Re: [ccp4bb] handling crystals in volatile solvents
Hi Len In every case that I know of this problem has been solved by working under oil. The oil becomes saturated with the volatiles, and it prevents crystals from doing backstroke, breaststroke or front crawl during harvesting. The most convenient way to get going is to dispense the wells without the volatile solvents, then allow solvents to difuse through the oil and into the drops. We suggest our old Vapor Batch plates, and you should place the same concentration of the solvent that you want in the drops into the large moat around the plate. If you put in higher concentrations you may dehydrate your drops :-( A very nice feature is that you can set up e.g. a random screen, then try soaking in one or more volatile solvents if nothing grows in the first round. The approach was invented by Lesley Haire, and you can find her presentation at http://www.douglas.co.uk/winner1.htm See also Mortuza, Gulnahar B., et al. High-resolution structure of a retroviral capsid hexameric amino-terminal domain. *Nature* 431.7007 (2004): 481-485. The plates look like this: http://www.douglas.co.uk/products.htm#Vapor Batch Plates We'll send you some sample plates to try. Best wishes, Patrick On 12 June 2015 at 22:11, Thomas, Leonard M. lmtho...@ou.edu wrote: Hi All, We have gotten some very nicely formed crystals out of a couple of different volatile solvents recently. Besides looking for something easier to work in does anybody have any tips on handling crystals from these types of solvents. It is very hard to loop a crystal while it is doing the backstroke in the well with all of its buddies. Thanks in advance. Len Leonard M. Thomas Ph.D. Macromolecular Crystallography Laboratory Manager University of Oklahoma Department of Chemistry and Biochemistry Stephenson Life Sciences Research Center 101 Stephenson Parkway Norman, OK 73019 405-325-1126 lmtho...@ou.edu http://barlywine.chem.ou.edu http://structuralbiology.ou.edu -- patr...@douglas.co.ukDouglas Instruments Ltd. Douglas House, East Garston, Hungerford, Berkshire, RG17 7HD, UK Directors: Peter Baldock, Patrick Shaw Stewart http://www.douglas.co.uk Tel: 44 (0) 148-864-9090US toll-free 1-877-225-2034 Regd. England 2177994, VAT Reg. GB 480 7371 36
Re: [ccp4bb] handling crystals in volatile solvents
This paper might be helpful. 1. J Struct Biol. 2014 Nov;188(2):102-6. doi: 10.1016/j.jsb.2014.09.011. Epub 2014 Oct 5. Efficient cryoprotection of macromolecular crystals using vapor diffusion of volatile alcohols. Farley C, Juers DH. Author information: (1)Department of Physics, Whitman College, Walla Walla, WA 99362, United States. (2)Department of Physics, Whitman College, Walla Walla, WA 99362, United States; Program in Biochemistry, Biophysics and Molecular Biology, Whitman College, Walla Walla, WA 99362, United States. Electronic address: juer...@whitman.edumailto:juer...@whitman.edu. Macromolecular X-ray crystallography, usually done at cryogenic temperature to limit radiation damage, often requires liquid cryoprotective soaking that can be labor intensive and damaging to crystals. Here we describe a method for cryoprotection that uses vapor diffusion of volatile cryoprotective agents into loop-mounted crystals. The crystal is mounted into a vial containing a small volume of an alcohol-based cryosolution. After a short incubation with the looped crystal sitting in the cryosolution vapor, the crystal is transferred directly from the vial into the cooling medium. Effective for several different protein crystals, the approach obviates the need for liquid soaking and opens up a heretofore underutilized class of cryoprotective agents for macromolecular crystallography. PMCID: PMC4252874 [Available on 2015-11-01] PMID: 25286441 [PubMed - in process] John J. Tanner Professor of Biochemistry and Chemistry Chair, Biochemistry Department Graduate Admissions Committee University of Missouri-Columbia 125 Chemistry Building Columbia, MO 65211 Phone: 573-884-1280 Fax: 573-882-2754 Email: tanne...@missouri.edumailto:tanne...@missouri.edu http://faculty.missouri.edu/~tannerjj/tannergroup/tanner.html On Jun 12, 2015, at 4:11 PM, Thomas, Leonard M. lmtho...@ou.edumailto:lmtho...@ou.edu wrote: Hi All, We have gotten some very nicely formed crystals out of a couple of different volatile solvents recently. Besides looking for something easier to work in does anybody have any tips on handling crystals from these types of solvents. It is very hard to loop a crystal while it is doing the backstroke in the well with all of its buddies. Thanks in advance. Len Leonard M. Thomas Ph.D. Macromolecular Crystallography Laboratory Manager University of Oklahoma Department of Chemistry and Biochemistry Stephenson Life Sciences Research Center 101 Stephenson Parkway Norman, OK 73019 405-325-1126 lmtho...@ou.edumailto:lmtho...@ou.edu http://barlywine.chem.ou.edu http://structuralbiology.ou.edu
[ccp4bb] handling crystals in volatile solvents
Hi All, We have gotten some very nicely formed crystals out of a couple of different volatile solvents recently. Besides looking for something easier to work in does anybody have any tips on handling crystals from these types of solvents. It is very hard to loop a crystal while it is doing the backstroke in the well with all of its buddies. Thanks in advance. Len Leonard M. Thomas Ph.D. Macromolecular Crystallography Laboratory Manager University of Oklahoma Department of Chemistry and Biochemistry Stephenson Life Sciences Research Center 101 Stephenson Parkway Norman, OK 73019 405-325-1126 lmtho...@ou.edu http://barlywine.chem.ou.edu http://structuralbiology.ou.edu
