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Thanks for the replies. I think I got it figured out. The pH seems to be the definitive factor. Indeed, the more alkaline the pH, the higher the amount of precipitation. I just played a bit around with the pH of Imidazole and it's cleared up. It also seemed to help to add things in this order..... Imidazole--->water--->ZnSO4--->PEG. Cheers, Chris On Tue, 2011-09-27 at 19:15 +0200, Enrico Stura wrote: > The PEG could also be the problem, so you can mix your stock solutions by > the method described > below and end up with the same result as soon as you add the PEG. > See: > Frances Jurnak: Effect of chemical impurities in polyethylene glycol on > macromolecular crystallization > Journal of Crystal Growth > Volume 76, Issue 3, 2 August 1986, Pages 577-582 > > Enrico. > > > > On Tue, 27 Sep 2011 18:43:09 +0200, Roger Rowlett <[log in to unmask]> > wrote: > > > Imidazole is basic. If you mix it directly with zinc salts you will get > > zinc > > hydroxide. Most screens are prepared by mixing stock solutions. In your > > case > > I would make up the screen from 1 M imidazole, pH 7.2, 1 M ZnSO4, and 50% > > PEG-4000. Add the PEG last. This should work. You may or may not need > > this > > specific salt or buffer to get crystals, and could swap them out or > > change > > concentration as required. 50 mM buffer may not be sufficient to control > > condition pH depending on the protein storage buffer composition. > > > > Roger Rowlett > > On Sep 27, 2011 11:56 AM, "Browning Christopher" < > > [log in to unmask]> wrote: > >> Dear All, > >> > >> My question might be a bit out of place, but perhaps someone can help. > > I've screened my protein in the Nuc-Pro screen from Molecular Dimensions > > and > > found some crystals in condition B10. They seem to be protein crystals as > > they are not highly optically active and look different to salt > > crystals. So > > condition B10 consist of 10% PEG 4K, 50mM Imidazole pH 7.2, 20mM Zinc > > sulfate. In the screen, this condition is perfectly clear, but when I try > > and make my own screen, the whole solution turns white. Apparently, Zinc > > sulfate/Imidazole can be used for staining SDS-gels, but that does not > > really help my a lot. Does anybody have an idea how I might be able to > > keep > > everything in solution, seeing that the MD guys managed somehow...... > > I'm a > > bit desperate as I don't have many hits for this protein. > >> > >> Cheers, > >> > >> Chris B > >> > >> -- > >> Dr. Christopher Browning > >> Post-Doctor to Prof. Petr Leiman > >> EPFL > >> BSP-416 > >> 1015 Lausanne > >> Tel: 0041 (0) 02 16 93 04 40 > > -- Dr. Christopher Browning Post-Doctor to Prof. Petr Leiman EPFL BSP-416 1015 Lausanne Switzerland Tel: 0041 (0) 02 16 93 04 40