Hi Morgan Elizabeth, Just following up on Vaheh’s suggestion, you can also add 2-4% of cryoprotectant, which includes your mother liquor (and your protein if crystals have dissolved), into the crystallization drop daily or weekly after crystals appear. Repeat this step until reaching the desired concentration of cryoprotectant. Additionally, I will harvest smaller crystals in your case.
If you can test the diffraction at room temperature, it seems to be the first priority for me to determine whether it is worthwhile to spend time on this crystallization condition and construct. Good luck! Best wishes, Tao-Hsin > On Nov 22, 2023, at 12:26 PM, Oganesyan, Vaheh > <[email protected]> wrote: > > Hi Morgan Elizabeth, > > In some cases adding 1-2% of cryoprotectant into crystallization drop during > setting those drops up helps to introduce 25-30% of the same cryoprotectant > during harvest, provided you still can get those crystals to grow. Worked for > me in several cases. > > Vaheh > > From: CCP4 bulletin board <[email protected] > <mailto:[email protected]>> On Behalf Of Blake, Morgan Elizabeth > Sent: Wednesday, November 22, 2023 11:44 AM > To: [email protected] <mailto:[email protected]> > Subject: [ccp4bb] Fragile Crystals > > Hello! > > I am a PhD student working on a crystallography project to wrap up my > dissertation research. I have purified a complex of two proteins, and I can > consistently grow crystals in 10% PEG3350, 0.2M KSCN, 0.1M BIS-TRIS propane > pH 7.5. These crystals have sharp edges and can grow to a large size (greater > than 0.5 mm), but the crystals seem to be very fragile. When we open the > drops to harvest the crystals, we have little time to harvest the crystals > before they crack. When we move the crystals to a cryoprotectant, over time > they start fracturing. We've tried using different percentages of glycerol, > ethylene glycol, PEG400, and oil for cryoprotectants with no success. > Needless to say, the crystals do not diffract well, with spot patterns that > look very streaky/mosaic, which I presume is due to the defects that we see > in harvesting/handling. We have screened for alternate crystallization > conditions, but we seem to get the same morphology in other conditions. Does > anyone have suggestions for additives we could use post-crystallization to > help stabilize our crystals? > > Thanks for your advice! > > To unsubscribe from the CCP4BB list, click the following link: > https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 > <https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1> > To unsubscribe from the CCP4BB list, click the following link: > https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 > <https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1> ######################################################################## To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list hosted by www.jiscmail.ac.uk, terms & conditions are available at https://www.jiscmail.ac.uk/policyandsecurity/
