I second Chris's suggestions. These have worked well for me in the
past. You only need a very thin layer of the grease (i.e. keep
wiping until its almost completely gone) and it usually has no affect
on the crystallization.
Jeff
On Jan 27, 2009, at 3:51 PM, Christopher Colbert wrote:
> If you have good and bad crystals in the same drop, I've had success
> pushing a crummy crystal into a good crystal and having it release
> that
> way.
>
> Additionally, once I realized this was going to be a long term
> problem, I
> started coating the sitting drop depressions with a thin layer of
> vacuum
> grease. The crystals just slid right off the grease and I never
> saw any
> changes in the diffraction data to suggest the grease was giving me
> issues.
>
> Chris
>
> On Wed, 28 Jan 2009, Savvas Savvides wrote:
>
>>>> Dear colleagues,
>>>>
>>>> we have been growing crystals of a protein complex in sitting-
>>>> drop geometry
>>>> that stick to the bottom of the drop remarkably well. It's as if
>>>> they are
>>>> glued onto the plastic. This makes crystal handling next to
>>>> impossible
>>>> without destroying the crystals. We have tried whiskers, loops,
>>>> all kinds of
>>>> micro-tools, and pipetting techniques to no avail. I can say at
>>>> the outset
>>>> that we have been unsuccessful in growing these crystals in
>>>> hanging-drops or
>>>> at 4 degrees. Deglycosylating the complex also leads to nowhere.
>>>> In fact, we
>>>> are only able to get crystals from homogeneously glycosylated
>>>> protein
>>>> produced in HEK293S/I- cells.
>>>>
>>>>
>>>>
>>>> In the meantime we are playing with the idea of siliconizing the
>>>> sitting-drop depressions to alter the crystal/plate interface.
>>>> But then
>>>> again, nucleation events on the plastic may be the reason we
>>>> are getting
>>>> crystals in the first place. We have also thought of trying
>>>> microseeding to
>>>> have more control on nucleation issues. Our protein production
>>>> is quite
>>>> limiting and forces us to be very selective with our
>>>> experimentation.
>>>>
>>>>
>>>>
>>>> Nonetheless, while we are waiting for fresh material to
>>>> explore some of
>>>> these ideas we would like to make the most out of the crystals
>>>> we have grown
>>>> thus far. We would therefore very much appreciate any input/
>>>> ideas on
>>>> manipulating these crystals for data collection.
>>>>
>>>>
>>>>
>>>> Best wishes
>>>>
>>>> Savvas
>>>>
>>>>
>>>>
>>>>
>>>>
>>>> ----
>>>> Savvas Savvides
>>>> L-ProBE, Unit for Structural Biology
>>>> Ghent University
>>>> K.L. Ledeganckstraat 35
>>>> 9000 Ghent, BELGIUM
>>>> office: +32-(0)9-264.51.24 ; mobile: +32-(0)472-92.85.19
>>>> Email: [log in to unmask]
>>>> http://www.lprobe.ugent.be/xray.html
>>>>
>>>>
>>>>
>>>>
>>>>
>>>>
>>>>
>>>> From: CCP4 bulletin board [mailto:[log in to unmask]] On
>>>> Behalf Of
>>>> Katarina Moravcevic
>>>> Sent: Tuesday, January 27, 2009 10:52 PM
>>>> To: [log in to unmask]
>>>> Subject: [ccp4bb] pseudo translation
>>>>
>>>>
>>>>
>>>> Hi all,
>>>>
>>>> here is a question from a beginner. I have a home source data
>>>> set that
>>>> indexed and scaled in a P2 space group (a=46.704,b=59.362,
>>>> c=48.783,
>>>> alpha=90.0, beta=104.469, gamma=90.0) with predicted 2mol/au.
>>>> After failing
>>>> to get a MR solution with Phaser I ran the phenix.xtriage which
>>>> showed that
>>>> I have a large (89.18) Patterson peak at 0.5 0 0.5 which
>>>> indicates pseudo
>>>> translational symmetry. I was wondering if there is anything I
>>>> could do with
>>>> this data to get around this problem. Given that I don't have a
>>>> lot of
>>>> experience any suggestion/explanation would be fantastic.
>>>>
>>>> Thanks in advance
>>>>
>>>> K
>>>>
>>>>
>>>>
>>>>
>>>>
>>>> E-mail message checked by Spyware Doctor (6.0.0.386)
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>>>>
>
> Christopher L. Colbert, Ph.D.
> Instructor Phone: (214) 645
> 5944
> University of Texas Southwestern Medical Center FAX: (214) 645
> 5945
> 6001 Forest Park Lane
> Dallas, TX 75390
>
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