Re: [ccp4bb] Detaching crystals from glass cover slides
Cool the coverslip on the opposite side of the crystal with a chip of dry ice. Do not freeze the drop. I learned this from Gary Gilliland. Also I wonder if you can simply move the whole tray into a cooler temperature? You can imagine that the thermal expansion coefficient of the glass coverslip and the crystal are different. I have not tried heating, but maybe that works, too? Or bend the coverslip without breaking is by applying pressure from the opposite side. Jim ... Any suggestions for detaching crystals from cover slides will be greatly appreciated. Wataru
Re: [ccp4bb] Detaching crystals from glass cover slides
Hi Wataru, I second Bill's suggestion. Additionally, you might want to migrate to a sitting drop setup and use the vacuum grease with that. It worked for me. Happy Fishing, Chris -- Christopher L. Colbert, Ph.D. Assistant Professor Department of Chemistry and Biochemistry North Dakota State University P.O. Box 6050 Dept. 2710 Fargo, ND 58108-6050 PH: (701) 231-7946 FAX: (701) 231-8324 On 2/8/11 8:19 PM, William G. Scott wgsc...@ucsc.edu wrote: Hi Wataru: I hope all is well. For the ones you already have grown, try very gently prying them off with a wedge-shaped needle. If you can grow more, try using a very thin smooth layer of vacuum grease, and apply the drop to that. I managed to get RNA crystals to grow that way that otherwise irreversibly adhered to the surface. All the best, Bill On Feb 8, 2011, at 6:06 PM, Wataru Kagawa wrote: Hi all, I have crystals growing by the hanging-drop method, using 24-well VDX plates and Hampton Research siliconized glass cover slides. Most crystals are attached to the cover slide, and I am having difficulties detaching the crystals (using a cryoloop) without breaking them. There are few smaller crystals floating in the drop, and they diffract X-rays pretty well (clean spots, ~3.5A resolution using RAXIS). However, I would like to try the bigger ones, because they may diffract to a higher resolution. Any suggestions for detaching crystals from cover slides will be greatly appreciated. Wataru
Re: [ccp4bb] Detaching crystals from glass cover slides
Hi Wataru, Have you tried Mitegen's polymer MicroTools? These tools have tips made from soft, flexible microfabricated polymer films and are specifically designed for this type of application. There is more information about them here ( http://mitegen.com/products/microtools/microtools.shtml). Best wishes, Ben Apker On Tue, Feb 8, 2011 at 9:06 PM, Wataru Kagawa wkag...@aoni.waseda.jpwrote: Hi all, I have crystals growing by the hanging-drop method, using 24-well VDX plates and Hampton Research siliconized glass cover slides. Most crystals are attached to the cover slide, and I am having difficulties detaching the crystals (using a cryoloop) without breaking them. There are few smaller crystals floating in the drop, and they diffract X-rays pretty well (clean spots, ~3.5A resolution using RAXIS). However, I would like to try the bigger ones, because they may diffract to a higher resolution. Any suggestions for detaching crystals from cover slides will be greatly appreciated. Wataru -- Product Development and Production Manager MiTeGen, LLC 95 Brown Rd - Suite 183 Ithaca, NY 14850 Phone: 607-266-8877 Fax: 607-697-0400 Disclaimer: The information contained in or attached to this e-mail is confidential, proprietary, and the sole property of MiTeGen, LLC. This e-mail is intended solely for the use of the intended recipient(s). If you have received this e-mail in error, please notify the sender immediately and then delete it. If you are not the intended recipient, you must not use, disclose or distribute this e-mail without the author's prior permission.
Re: [ccp4bb] Detaching crystals from glass cover slides
Dear Wataru check out the following entry on the CCP4-wiki on 'sticky crystals' which we compiled based on feeback from many crystallographers: http://strucbio.biologie.uni-konstanz.de/ccp4wiki/index.php/Sticky_crystals Best regards Savvas Savvas Savvides Unit for Structural Biology @ L-ProBE Ghent University K.L. Ledeganckstraat 35, 9000 Ghent, Belgium Tel/SMS/texting +32 (0)472 928 519 Skype: savvas.savvides_skype http://www.LProBE.ugent.be/xray.html On 09 Feb 2011, at 16:11, Colbert, Christopher wrote: Hi Wataru, I second Bill's suggestion. Additionally, you might want to migrate to a sitting drop setup and use the vacuum grease with that. It worked for me. Happy Fishing, Chris -- Christopher L. Colbert, Ph.D. Assistant Professor Department of Chemistry and Biochemistry North Dakota State University P.O. Box 6050 Dept. 2710 Fargo, ND 58108-6050 PH: (701) 231-7946 FAX: (701) 231-8324 On 2/8/11 8:19 PM, William G. Scott wgsc...@ucsc.edu wrote: Hi Wataru: I hope all is well. For the ones you already have grown, try very gently prying them off with a wedge-shaped needle. If you can grow more, try using a very thin smooth layer of vacuum grease, and apply the drop to that. I managed to get RNA crystals to grow that way that otherwise irreversibly adhered to the surface. All the best, Bill On Feb 8, 2011, at 6:06 PM, Wataru Kagawa wrote: Hi all, I have crystals growing by the hanging-drop method, using 24-well VDX plates and Hampton Research siliconized glass cover slides. Most crystals are attached to the cover slide, and I am having difficulties detaching the crystals (using a cryoloop) without breaking them. There are few smaller crystals floating in the drop, and they diffract X-rays pretty well (clean spots, ~3.5A resolution using RAXIS). However, I would like to try the bigger ones, because they may diffract to a higher resolution. Any suggestions for detaching crystals from cover slides will be greatly appreciated. Wataru
[ccp4bb] Detaching crystals from glass cover slides
Hi all, I have crystals growing by the hanging-drop method, using 24-well VDX plates and Hampton Research siliconized glass cover slides. Most crystals are attached to the cover slide, and I am having difficulties detaching the crystals (using a cryoloop) without breaking them. There are few smaller crystals floating in the drop, and they diffract X-rays pretty well (clean spots, ~3.5A resolution using RAXIS). However, I would like to try the bigger ones, because they may diffract to a higher resolution. Any suggestions for detaching crystals from cover slides will be greatly appreciated. Wataru
Re: [ccp4bb] Detaching crystals from glass cover slides
Hi Wataru: I hope all is well. For the ones you already have grown, try very gently prying them off with a wedge-shaped needle. If you can grow more, try using a very thin smooth layer of vacuum grease, and apply the drop to that. I managed to get RNA crystals to grow that way that otherwise irreversibly adhered to the surface. All the best, Bill On Feb 8, 2011, at 6:06 PM, Wataru Kagawa wrote: Hi all, I have crystals growing by the hanging-drop method, using 24-well VDX plates and Hampton Research siliconized glass cover slides. Most crystals are attached to the cover slide, and I am having difficulties detaching the crystals (using a cryoloop) without breaking them. There are few smaller crystals floating in the drop, and they diffract X-rays pretty well (clean spots, ~3.5A resolution using RAXIS). However, I would like to try the bigger ones, because they may diffract to a higher resolution. Any suggestions for detaching crystals from cover slides will be greatly appreciated. Wataru
Re: [ccp4bb] Detaching crystals from glass cover slides
Dear Wataru-san: I understand the problem and sometimes I also faced the same difficulties. One likely suggestion to pick up the crystal is here: 1. Hold the cover-glass in hand (the drop should be facing bottom side as in the hanging drop setup) 2. Adjust the microscope to see the crystals (bottomside) and decide which one you are going to pick-up (keep in mind that you are going to use cryo-loop, so you need enough space between the cover glass and bottom of the microscope) [please look at the attachements for quick understanding] 3. Use the cryoloop from the bottom of the coverglass and pick the crystals. moving the crystals might be easier... all the best. With regards, Kumarevel -Original Message- From: CCP4 bulletin board [mailto:CCP4BB@jiscmail.ac.uk] On Behalf Of Wataru Kagawa Sent: Wednesday, February 09, 2011 11:06 AM To: CCP4BB@jiscmail.ac.uk Subject: [ccp4bb] Detaching crystals from glass cover slides Hi all, I have crystals growing by the hanging-drop method, using 24-well VDX plates and Hampton Research siliconized glass cover slides. Most crystals are attached to the cover slide, and I am having difficulties detaching the crystals (using a cryoloop) without breaking them. There are few smaller crystals floating in the drop, and they diffract X-rays pretty well (clean spots, ~3.5A resolution using RAXIS). However, I would like to try the bigger ones, because they may diffract to a higher resolution. Any suggestions for detaching crystals from cover slides will be greatly appreciated. Wataru cryoloop.ppt Description: MS-Powerpoint presentation
Re: [ccp4bb] Detaching crystals from glass cover slides
Have you tried seeding ? And another suggestion, if the small ones already diffract to 3.5 Å on a home source, why don't you try to go to a synchrotron ? Big is also not always better, they might freeze worse than your small ones or might have growth artifacts etc. If you take several huge crystals and touch them gently with e.g. a hair from a Unicorn* then let it sit and recover you might get lucky. Jürgen * can be replaced by any stronger hair of your preference e.g. pot-bellied pig, horse, cats anything that's a bit more sturdy or simply the Microtools from HR - Jürgen Bosch Johns Hopkins Bloomberg School of Public Health Department of Biochemistry Molecular Biology Johns Hopkins Malaria Research Institute 615 North Wolfe Street, W8708 Baltimore, MD 21205 Phone: +1-410-614-4742 Lab: +1-410-614-4894 Fax: +1-410-955-2926 http://web.mac.com/bosch_lab/http://web.me.com/bosch_lab/ On Feb 8, 2011, at 9:06 PM, Wataru Kagawa wrote: Hi all, I have crystals growing by the hanging-drop method, using 24-well VDX plates and Hampton Research siliconized glass cover slides. Most crystals are attached to the cover slide, and I am having difficulties detaching the crystals (using a cryoloop) without breaking them. There are few smaller crystals floating in the drop, and they diffract X-rays pretty well (clean spots, ~3.5A resolution using RAXIS). However, I would like to try the bigger ones, because they may diffract to a higher resolution. Any suggestions for detaching crystals from cover slides will be greatly appreciated. Wataru
