Re: [ccp4bb] Intergrown crystals and excess of nucleation

[Claude Sauter]

Le 09/09/2019 à 17:22, Nikolas a écrit : Dear crystalgrowers, I am currently working with a protein that appeared to be friendly but turned out it was not the case. I found myself to face -in the scale up- the opposite of the usual problem of nucleation (I really love how this topic finds new ways to make fun of me). In 24-well plates, hanging-drop, for the same condition but in different drops I found few big but intergrown crystals and/or a full with microcrystals. Sometimes also in the same well, when having more drops. I already decreased the concentration to less than 4mg/mL, made small adjustments in the optimizations - both with apo and ligand samples, used Al's oil.  I have read about the "containerless crystallization" but since I cannot obtain the sample myself I would like to know if there are any experiences and/or if there are suggestions for solving this problem. Many thanks! Best regards, Nikolas To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB&A=1 Dear Nikolas, since you have some crystal stock, I would definitely try seeding to better control nucleation events in your drops. Then, instead of using the containerless approach which requires two types of oils to prepare floating drops, I suggest the crystallization in agarose gel. Easy to perform, it favors the 3D growth of well separated crystals in ideal convection-less conditions. In addition, the gel provides a physical protection of your crystals during handling, mounting and cryocooling. For more details, see "Crystal growth of proteins, nucleic acids, and viruses in gels. Lorber et al. Prog. Biophys. Mol. Biol. (2009), 101: 13-25." Happy crystallization! Claude -- Dr Claude Sauter Institut de Biologie Moléculaire et Cellulaire (IBMC-ARN-CNRS) Biologie des ARNt et pathogénicité tel +33 (0)388 417 102 2 allée Conrad Roentgen fax +33 (0)388 602 218 F-67084 Strasbourg - France http://cj.sauter.free.fr/xtal To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB&A=1