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Figure 1
The CrystalDirect approach. (a) Schematic representation of the methods for automated crystal harvesting, chemical delivery and cryocooling. From top to bottom, crystals are grown on the surface of a low X-ray-background film which is directly compatible with diffraction data collection. A laser beam operating in the photoablation regime is used to produce an aperture in the film (1). Chemicals can be delivered to crystals through diffusion by applying a small amount of solution on the outside of the opening that enters into contact with the crystallization drop (2). After incubation, or immediately after producing the aperture, if no chemicals are delivered both the externally applied and the crystallization solution are gently aspirated through the aperture by applying a vacuum (3–4). The sample can then be mounted by excising the film around the crystal with the laser and gluing it to the tip of a data-collection pin (5). The crystal is then moved to a cryojet for flash-cooling (6). (b) The 96-well CrystalDirect microplate. One of the cells of the microplate is shown in detail (outlined in red). The reservoir containing the crystallization solution (left) and the film used as the crystallization support (right) with three crystallization drops on it can be appreciated. Crystals from one of the drops have been harvested and mounted on a pin (blue outline). (c) X-ray diffraction analysis of crystals of proteinase K (left), thaumatin (middle) and lysozyme (left) prepared using the automated CrystalDirect harvesting and cryocooling method. The top panels show a detail of the mounted samples as seen through the on-axis beamline camera (the blue circle indicates the size and position of the X-ray beam). Only a small amount of liquid remains between the film and the crystal after the liquid-removal step, facilitating direct cryocooling. X-ray diffraction images from these samples (middle panels) show a complete absence of ice rings and show reflections extending to high resolution with spot profiles (bottom panels) and crystallographic statistics comparable to those obtained with crystals processed with the standard manual methods (see Table 1[link]).

Journal logoSTRUCTURAL
ISSN: 2059-7983
Volume 72| Part 4| April 2016| Pages 454-466
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