re : Paratone-N oil.
1) I find it much easier to work with if you cut it 50:50 with light
mineral oil.
2) he stress of moving mechanically weak crystals (needles or thin
plates) from a crystallisation liquor to the more viscous Paratone can
cause the crystal to visibly bend - which might not be such a good
thing. But I find that paratone works really well for most chunky,
solid crystals.
re: the high mosiacity issues - maybe you could try some gentle
dehydration of the crystals? Maybe preincubate the crystals in mother
liquor + 10>20% PEG20k prior to freezing?
HTH
Dave
2009/10/7 Janet Newman <[log in to unmask]>:
> One thing to do here is to work out where you have your problem.
>
> Is the crystal mosaic after you have introduced your potential cryo solutions, but still at room temperature? If it is, there is very little chance (snowballs in hell type chance) that it will get less mosaic when you flash cool it.
>
> Can you transfer your crystal from the high AmPO4 to high AmSO4 ? or LiSO4 or malonate (all known cryo-salts) and flash cool from there?
>
> Janet
> ________________________________________
> From: CCP4 bulletin board [[log in to unmask]] On Behalf Of ycheng [[log in to unmask]]
> Sent: 08 October 2009 07:54
> To: [log in to unmask]
> Subject: [ccp4bb] Questions about cryoprotectant
>
> 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
>
--
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Science knows it doesn't know everything...otherwise it would stop.
- Dara O'Briain
============================
David C. Briggs PhD
Father & Crystallographer
http://drdavidcbriggs.googlepages.com/home
Skype: DocDCB
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