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Subsections


Screening Crystals

The Screening tab provides an interface to view the results of automated screening with the Blu-Ice/DCS software. The screening tab allows viewing and editing of the spreadsheet containing sample information and diffraction analysis results and also detailed inspection of the images and results obtained for each screened sample.

When clicking on the Screening Tab for the first time in the session, the browser displays the User Cassettes, a list of the spreadsheets previously uploaded to SSRL by the user. When a beamline is selected, the Beamline Cassettes will list the spreadsheets currently assigned to each cassette position in the beamline.

The search tool at the top makes it easy to locate a specific spreadsheet. The search input box allows use of the wild-card ''*''; in this case all the spreadsheets matching the partial name will be displayed.

The spreadsheet contents are accessed by clicking on one of the following links:

  • Summary: Displays the selected spreadsheet in a table format. It also allows access to a tool to edit the spreadsheet contents.

  • Details: Allows detailed inspection of the diffraction image analysis and autoindex results for each sample.

  • Download Excel: Allows the user to download the spreadsheet to the user's computer. If the spreadsheet has been used in the automatic screening, the image analysis and autoindexing results will be in the spreadsheet.

The link Upload new spreadsheet directs to the Screening System Database (usually to upload a new spreadsheet).

Figure 25: Spreadsheet list with links to the screening results, Excel spreadsheet file and the Screening Database interface.
Image screen-all-cassettes

Once a cassette has been selected, it is possible to revisit the results for that cassette by clicking on the Cassette Summary and Cassette Details links in the Screening Tab navigation toolbar.


Cassette Summary

In the Cassette Summary view, the spreadsheet is displayed in a table format. All rows (each corresponding to a sample) are displayed. The columns and the contents displayed can be controlled by the display options set in the Preferences Tab.

Figure 26: Cassette Summary: the users' spreadsheet is displayed in table format.
Image screen-cassette-summary


Screening Results

If the user collects two images during screening, autoindexing of the images will be carried out automatically. The following columns will be shown in the Cassette Summary - Results view and appended to the user's spreadsheet:

  • Unit cell and Bravais lattice. Note that the symmetry is, at this point just an educated guess. The actual Laue group and the presence of pseudo-symmetry can only be detected during scaling. Sometimes, particularly if the diffraction is poor, a lower symmetry than the correct one can be chosen by LABELIT.

  • Mosaicity: The initial mosaicity estimate by MOSFLM. The estimate is based on how many observed spots are predicted and it is good enough for strategy calculation and sample characterization. A more accurate value can be obtained after post-refinement using a larger number of images.

  • Rmsr: Root mean square deviation between the observed and predicted spots, as calculated by MOSFLM after image integration. It is usually around 0.05 or less for good crystals.

  • Resolution: The diffraction limit of the crystal at the used exposure time based on the $I/\sigma I$ values. The limit is set at $I/\sigma I = 1.5$ If the diffraction is very strong, the resolution may extend beyond the limit set by the detector area and sample to detector distance used during screening.

  • Score: Calculated as $1.0 - (0.7 \times e^{-4/d}) - (1.5 \times rmsr) - (0.2 \times \mu)$, where $d$ is the resolution limit in Å, rmsr (described above) is in mm, and $\mu$ is the mosaicity in degrees. A score of 0.4-0.6 indicates borderline quality. Good crystals usually have a score of about 0.8 or higher. This score is meant to be used as an initial indicator of the sample quality; although it has been found to agree reasonably well with human assessment in many cases, it is recommended to inspect the diffraction before choosing a sample for data collection.

  • Image: Diffraction image name.

  • System Message: If the autoindexing fails, the error message from LABELIT will be displayed in this column.

A second set of results are appended to the Excel file as a separate column for each diffraction image, regardless of whether autoindexing has been performed or not. In the Cassette Summary page they can be seen by going to the Preferences Tab and setting the Image Display Type set to ''Show all Crystal Links'' (for all samples) or ''Show Selected Images'' (only for one sample). If a group of two or more contiguous images are collected at each crystal orientation, results are given only for the last image in each group.

These values are particularly useful if the autoindex fails. Having an independent quality assessment for each image is then useful to determine if the sample merits further inspection or it can be safely discarded.

  • Crystal JPEG: Snapshot file name.

  • Spot Shape: Mean value of the longest spot dimension divided by the shortest one. It is between 1.5 and 2 for good crystals.

  • DISTIL Resolution: Resolution estimate by the LABELIT SPOTFINDER module, based on the number of spots in resolution shells.

  • Ice rings: Number of ice rings.

  • Diffraction Strength: Average spot intensity.

  • Score: The score for the individual image. 0 or 1 usually indicate no useful diffraction. A sample scoring higher than 4-5 may merit inspection. Very different scores for images at different orientation indicate anisotropic diffraction or a badly centered crystal; this can be confirmed by inspection of the Crystal JPEG.

Selecting samples

A sample is selected by clicking the radio button in the port column; only one button can be selected at a time.

Figure 27: Selecting the sample row A1.
Image screen-port-select


Data collection strategy

A data collection strategy will be calculated during screening if the sample stays mounted on the goniometer head3 after the screening shots (i.e., by inserting a ''Stop'' in the screening task sequence in Blu-Ice - see the Blu-Ice User Manual). In this case, the View Strategy button will direct the user to the Autoindex Tab, from which the strategy results can be viewed once the strategy calculation is complete. If the strategy calculation has not been finished, the Update button in the Autoindex Tab to update the contents of the run.

Re-analyzing samples

The Analyze Crystal button can be used to re-run the analysis and autoindexing software for a selected sample. This is useful if the test images for that sample have been recollected or in case of a software crash or server time-out during the first analysis.

While a sample is being re-analyzed the Analyze Crystal button will be disabled. Use the Update button to display the new analysis results.

Editing spreadsheet columns

To access the sample editing tool, select a crystal as described above; then, click the Edit crystal button at the top of the spreadsheet table. This will link to the the Edit crystal tool page (shown in figure 28). The selected sample row is displayed and only certain fields are editable. Click Set Crystal to save the changes. To cancel the changes, simply exit the page by clicking the Cassette Summary button (to return to the spreadsheet table) or All Cassettes or Cassette Details to access the user's spreadsheet list or the detailed screening result viewer respectively.

Figure 28: Sample editing tool
Image screen-edit-sample

Sorting samples

Some column headers are displayed as links. Clicking on will result in column sorting, toggled between ascending and descending. For example, to sort the samples by score, click on the Score column header.

Note: The sorting affects the table display only. The actual Excel spreadsheet is always sorted by port ID unless the user edits the Excel file directly.


Updating the Spreadsheet

The spreadsheet is updated with the latest available screening results automatically every 5 seconds by default; this can be changed from the Preferences Tab.

The Update button can also be used to reload the spreadsheet from the crystals server.


Cassette Details

The Details page is similar to the Image Viewer tab; but instead of using the file browser to locate the image file, the user uses the cassette browser.

The top left frame displays one of the images of the selected crystal. Like in the Image Viewer, the user can zoom, pan, and adjust the brightness. The Analyze Image button displays the image with highlighted spots picked by SPOTFINDER. The user can toggle between Analyze Image and Image Only to display or hide the highlighted spots.

The top right frame contains various tabs that display information about the image and the autoindexing results:

  • Header: The Header tab displays the image header.

  • Analyze Image: This tab displays the image analysis results. The analysis results (logs and JPEG images) are stored together with the user's data under the directory crystal-analysis.

  • Crystal Image: The Crystal Image tab displays the crystal snapshot JPEG file.

  • Autoindex: This tab displays the autoindex results from LABELIT. Autoindex will be carried out if two images at different crystal orientations are collected. Image or images collected at a third orientation will not be used for the autoindex.

  • Details: The Details tab lists all the output files from autoindexing. The results are stored together with the user's data in the directory listed at the top of the Details view. The files named indexXX and indexXX_S. mat are the MOSFLM input script and matrix used for solution number XX. By clicking on a file name, the contents will be displayed on a separate browser window.

The bottom frame contains the cassette browser tool. It displays an array corresponding to the ports in the cassette (or unipuck). The ports that contain valid crystal id are displayed with links, otherwise they are grayed out. When the user clicks one of the ports, the first image collected for that port will be displayed.

The Analyze and View Strategy buttons have the same functionality as in the Cassette Summary page.

Note: The Update function and button only activate the link to the latest sample for which results are available. It does not update the images and results displayed.


Footnotes

... head3
Typically, the crystal orientation will be different after each sample remount, so the collection strategy will change accordingly.