Hi Everyone,
Franks point is really interesting. We routinely reduce the protein
concentration when we see too many precipitated wells, but we never
dilute the screen. Has anyone tried this?
All the best,
Alun
On 12/07/17 08:48, Frank von Delft wrote:
The point I was failing to make: reducing either protein or
precipitant concentration will indeed reduce nucleation, but often
won't get you bigger or more single crystals: it will just make the
appearance of crystals less reliable.
The way to get big single reliable crystals is to /increase/ protein
and /greatly/ reduce precipitant.
(Even better: do seeding. Like Vicky said. Incredible how often
people don't bother to do seeding, yet it solves so many problems.)
phx
On 12/07/2017 07:50, Vicky Tsirkone wrote:
Dear Frank,
I may see in the attached pic several nucleation points and a
considerable amount of microcrystals. Based to my knowledge
decreasing the concentration of the precipitant and/or the protein
concentration would be a reasonable approach to refine the initial hits.
By checking the diagram as you correctly mentioned you may see that
the fine tuning of protein and precipitant concetration may lead to
the desirable result without reaching the precipitation zone.
Patrick just check your screens. Just a rule of thumb, if you see
precipitation in the ~60% of your drops then you should definitely
reduce the protein concentration.
ps dont forget to try the *streak seeding*, as well.
Have a nice day and again good luck.
Vicky
On Wed, Jul 12, 2017 at 8:50 AM, Frank von Delft
<frank.vonde...@sgc.ox.ac.uk <mailto:frank.vonde...@sgc.ox.ac.uk>> wrote:
Actually, you should try /increasing/ the protein concentration -
a lot. But be prepared to drop the precipitant concentration to
almost nothing (1 or 2% isn't "low").
To understand why, look at the phase diagram and what we assume
about vapour diffusion. (Which I'm assuming is what you're doing.)
On 12/07/2017 06:28, Vicky Tsirkone wrote:
Dear Patrick,
You may reduce the protein concentation, as well.
Another option could be the *streak seeding* by exploiting the
drop of your initial condition.
Good luck,
V.T.
On Mon, Jul 10, 2017 at 7:17 PM, Patrick Shaw Stewart
<patr...@douglas.co.uk <mailto:patr...@douglas.co.uk>> wrote:
Microseed them into two or three random screens.
Search for MMS and rMMS online.
Good luck
Patrick
On 10 July 2017 at 15:47, Liuqing Chen <519198...@163.com
<mailto:519198...@163.com>> wrote:
hello everyone!
I get a condition (10% w/v PEG 6000, 100mm HEPES PH7.0)
in which my protein grow small needle like crystals,
how can i optimize it to get bigger crystals? the
attach is the crystals figure.
thanks in advance
sincerely
Liuqing Chen
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