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The following sections describe how to carry out an experiment at from sample screening to full data collection making use of the integrated data collection and analysis environment at SSRL.
For additional help setting up data collection, please consult the Blu-Ice documentation.
The high throughput screening system implemented at SSRL makes it possible to automatically collect and analyze test images and fully characterize the sample in a semi- or fully automated fashion.
A summary of the autoindexing results (symmetry, resolution and mosaicity estimates and a score) will be written to the sample list information displayed in Blu-Ice and Web-Ice shortly after the images have been collected. This information can be used to help select the best samples for collection of a complete data set. Follow the instructions below or consult the Video Tutorial
To see the screening results in the Blu-Ice screening tab, make sure that you use the ''Results''view to see these columns.
You can inspect the results in detail with Web-Ice:
Once the optimal sample for data collection has been selected, monochromatic (non-anomalous), and simple MAD and SAD experiments from a single crystal can be set up in a very easy way using Web-Ice. Follow the steps below or look at the Video Tutorial
For monochromatic experiments the priority is to maximize unique completeness. If the estimated dose for the experiment is low enough, consider collecting additional data beyond the starting or ending phi to increase data redundancy.
Completeness for the low resolution shells is important, so decrease the exposure time or use additional attenuation if the strategy page displays a warning about overloaded spots in one or both of the images.
If ultra-high resolution data are required and you are on a beamline equipped with a CCD detector (e.g. BL14-1) it may not be possible to collect to the resolution limit without overloading the low resolution reflections. In this case, collect an additional low-resolution pass:
A low pass may not be necessary when collecting thin phi sliced data on the high dynamic range Pilatus detector, but check the reflections at low resolution.
For MAD and SAD experiments is it very important to limit the dose received by the crystal during the experiment. The Web-Ice strategy already incorporates some mitigation procedures (e.g., data collection in wedges, use of two wavelengths for MAD experiments). If the Web-Ice strategy still results in a dose exceeding or at the limit given by the software, consider decreasing the exposure time. This can be done manually either after exporting the data collection parameters to Blu-Ice, or in the edit window Web-Ice displays prior to initiating data collection. The dose is proportional to exposure time, so reducing the exposure time by half will reduce the dose by half at the only expense of a slightly lower data resolution.
For certain space groups and crystal orientations, it is also possible to reduce the absorbed dose by selecting the phi range to maximize unique data set completeness (maximizing Bijvoet pair completeness is the default strategy for MAD and SAD). This usually works for MAD data with a medium to strong anomalous signal.
In unfavorable cases (very small, weakly diffracting crystals) it may be impossible to collect a data set without inflicting serious radiation damage to the crystal. In this case, several crystal will most likely be required for successful structure solution.
Low signal experiments
If the expected anomalous signal is very low (less that 1 % of the average reflection intensity), collection of additional redundancy may be required for structure solution. This can be done by manually adjusting the ending phi before proceeding with data collection in Blu-Ice or Web-Ice. As mentioned above, the exposure time may also require adjusting to avoid excessive irradiation of the crystal.
On the most intense beamlines, particularly BL12-2, it is possible to collect data from small crystals of dimensions less than 20 microns. However, because diffracted intensity decreases with the crystal volume faster than the deposited dose, use of such small crystals often prevents being able to collect a full data set from a single crystal before radiation damage severely affects the data. This is also often the case for data collection at above cryo and room temperatures, were often crystals last about a factor of 100 less than at cryo. In this cases, it is necessary to stitch a data set from data collected from different crystals. For high symmetry space groups it is often possible to obtain good completeness by starting data collection on a random orientation for each crystal. For low symmetry or scarce samples, it is useful to determine a data collection strategy for each crystal that maximizes the total completeness. This can be achieved with the collection strategy tool available in Web-ice.
Some tips for multicrystal experiments:
The following directories are automatically created the first time you log in to a SSRL px computer (these directories are accessible from all computers):
Use the remote data processing servers (pxproc09 - pxproc32) to process the data; from the beamline workstations, use the SSRL menu option of the Xfce panel in the Linux beamline computers and the remote Unix desktop. You can also right-click on the Linux Xfce desktop and select Data Processing from the desktop menu. Clicking on Select Least Loaded displays the load. Avoid using a computer is the load is close to the total number of CPUs (displayed next to the computer name).
Note that, since 2017, Web-Ice attempts automated data processing after a data set has been collected. The purpose of this tool is to provide quick feedback of the data quality. Please consult the Web-Ice data processing help.
more information about the beamline machines, consult the web document
Commonly used software packages for data processing are available at the macromolecular crystallography beamlines. If you are unfamiliar with a particular application, consult the relevant documentation:
For a complete list of supported and unsupported software installed in the
SSRL computers, see
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questions: Ana Gonzalez
|Last modified:Wednesday, 23-May-2018 16:49:50 PDT.|