I find it unlikely that you are seeing PEG400 in the solvent. I have a glycosylated protein, data collected at 100 deg, and can only make out the first one or two sugars, and these are covalently attached to the protein! To overcome the low occupancy problem, the first thing I would try is co-crystallization. I had been using glycerol as a cryoprotectent, then ran into problems with some new crystals (the crystals cracked, no matter how I did the soak). I tried using no cryoprotectant instead: I just dropped the crystal into a very heavy mineral oil, then lightly stirred the oil around the crystal to remove mother liquor from the surface of the crystal. I froze the crystal in liquid N2, and transferred to the stream. The crystal diffracted nicely. There were no ice rings, and an "oil ring" instead of a water ring, and the data seem fine (no problems with processing). Other people in our lab have had good experiences with this technique as well. ******************************************************************************** Brian Shilton Phone: (514) 496-6784 National Research Council Canada Fax: (514) 496-5143 Biotechnology Research Institute Email: brian.shilton@bri.nrc.ca 6100 Royalmount Ave. Montreal, Quebec H4P 2R2 Canada ********************************************************************************
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