you may have thought of this already, but you could try cryoprotection in the drop itself. i.e. slowly adding cryoprotectant to the reservoir, or replacing the reservoir bit by bit with solution containing cryoprotectant, and then adding small volumes to the side of the drop - for example, exchanging 20% volume cryo-solution with the reservoir, letting equilibrate with the unchanged drop for a few hours, then add 20% volume to the drop from the reservoir (i.e. 0.2 ul if the drop is 1 ul), then add another 20% of cryo to the reservoir, equilibrate a few hrs, etc. - the idea being to very slowly change the drop conditions and minimise risk of cracking. Of course, you may need patience and many drops of crystals, not just many crystals in a few drops, until you find the cryoprotectant where the crystals do not crack and still diffract, if you are successful at all... but if it works, you can just harvest from the equilibrated drop and directly flash-cool
Mark J van Raaij Laboratorio M-4 Dpto de Estructura de Macromoleculas Centro Nacional de Biotecnologia - CSIC c/Darwin 3 E-28049 Madrid, Spain tel. (+34) 91 585 4616 http://www.cnb.csic.es/content/research/macromolecular/mvraaij On 26 Oct 2011, at 18:46, Leonard Thomas wrote: > Hi All, > > I have run into a very sensitive crystals system when it comes to cryo > protecting them. I have run through the usual suspects and trays are going > to be setup with a cryo protectant as part of crystallization cocktail. The > one problem that seems to be occurring is that the crystals crack as soon as > they are transfered out of the original drop. I am running out of ideas and > really would love some new ones. > > Thanks in advance. > > Len > > Leonard Thomas Ph.D. > Macromolecular Crystallography Laboratory Manager > University of Oklahoma > Department of Chemistry and Biochemistry > Stephenson Life Sciences Research Center > 101 Stephenson Parkway > Norman, OK 73019-5251 > > lmtho...@ou.edu > http://barlywine.chem.ou.edu > Office: (405)325-1126 > Lab: (405)325-7571
