You probably included images when creating a new project. The Quality Control step allows you to view these images and their properties, check their quality and consistency, edit them when necessary and define pI and MW markers for automatic pI/MW calibration. You can also add, remove or rename images. When finished, you can validate those gels you want to take forward for further analysis.
Most of the functionality in this step is specifically carried out on selected images. You can select images by clicking on their thumbnail. To select multiple images, hold the Ctrl or Shift keys.
Note the difference between images and gels: gels are the physical objects for which one or several images have been acquired. In the case of DIGE and other multi-stain experiments, two or more images belong to the same gel.
To rename images or gels, click on their name above the thumbnail or in the table.
By default, the Quality control screen displays the thumbnails for all imported gel images, with below, the Quality control table. The table shows various properties of the gel images: Name, image height and width in pixels, bit depth, estimated bit depth, number of gray levels used, minimum and maximum gray levels before and after calibration, the percentage of gray levels used (calculated with respect to estimated bit depth), the percentage of the dynamic range in use (calculated with respect to estimated bit depth), the percentage saturated areas and background clipping detected, as well as the calibration formula and unit.
You can change your preferred way of viewing images and image information in the Quality control screen.
- Images/Table – Display both the image thumbnails and the quality control table.
- Images – Only display the image thumbnails, and hide the quality control table.
- Table/Histogram – Display the quality control table with the gel image and intensity histogram columns included.
Potentially problematic items, issues or values are highlighted at various levels in the Quality control screen, through icons and color codes:
- Pass – green – All looks correct, you can confidently proceed with your analysis.
- Caution – yellow – Acceptable, but be careful, Melanie detected an issues that could weaken the quality of the analysis.
- Warning – red – Anomaly identified, Melanie detected an issue that will adversely affect the quality of the analysis.
Click the More icon to display detailed information about the detected issue, how it could affect the analysis, and what you can do to solve it.
The Edit images button will take you to a dedicated Image edition mode that provides the basic tools to edit images:
(Only images that were selected in the Quality control step are opened in the Image edition mode.)
- Rotate left (90° CCW)
- Rotate right (90° CW)
- Rotate free
- Flip horizontally
- Flip vertically
- Invert gray levels
- Change resolution
Note that editing always occurs on a copy of the imported image file; never on the original file itself.
In the Image edition mode, you will also be able to display a 3D view of your images, and to adjust their contrast. These two display options do not alter the underlying image data.
Choose Edit pI/MW from the Edit button drop down to create and edit pI and MW markers on known proteins. Melanie uses the pI and MW values of these proteins to compute approximate pI and MW values for all spots. The pI/MW information will automatically be available in all images and reports throughout the analysis.
In the Edit pI/MW screen, first choose the image on which you want to define the markers. You can define markers on several images. But only markers on the image currently selected in the Image for pI/MW box will be used to calculate theoretical values for all other spots. Note that in the main Quality control screen, the image used for pI/MW calibration is marked with a small pI/MW icon.
Click the Edit pI/MW icon to enter the corresponding mode, and then double-click the position where you want to create a pI or MW marker. In the Create pI/MW marker box, specify whether you want to create a pI or MW marker and enter its value.
- To edit a marker’s value, double-click its label.
- To move a marker’s position, drag its base (disc).
- To delete a marker, right-click on its base (disc). Right-click and drag to delete all markers in an area.
Click the icons in the Display section to access helpful tools:
- Adjust contrast – Allows you to adjust the contrast of your image.
- 3D view – Shows the 3D view, to help more precise positioning of your markers.
- Grid – Displays the pI and/or MW grid on your image.
The pI/MW table lists the markers that have been defined, with their X and Y coordinates.
You can systematically review each gel to decide whether you want to take it forward for further analysis.
Your choice will be reflected in the Validation box at the top right of each gel image, and at the left of the gel or image name in the Quality control table. You can also click this box to choose between the three different states:
- The gel will be included in the analysis.
- The gel will be excluded from the analysis.
- the gel has not been validated, but will be included in the analysis by default.