This page describes the changes and fixes that were introduced in the Melanie software since the release of Melanie 8.0.0 (2015.12.23.0).

You can learn about the installation requirements and instructions for Melanie 8.0.1 (2016.7.6.0) in its Readme file.

Changes

Quality control

• On first launch of the software, the default path for opening image files points to the folder with tutorial images.
• Opening image or software-related files (.prj, .mel, .gda, .mda) by drag and drop onto the program window or by using Open with has been forbidden to avoid project corruption.
• HTML in the left feedback panel has been improved.
• Calculation of Estimated bit depth has been refined.

Experimental design

• Improved management of cases where no or only one level is defined for a factor.
• More precise information is provided when a design is not supported or not ideal.
• Column widths are adapted to the length of the factor names.

Alignment

• Improvement and optimization of alignment.
• Number and percentage of matches are now always visible, regardless of the length of the image names.

Detection

• Minor improvements in Detection parameters dialog to allow better fine-tuning of parameters, and fit larger parameter values.

Review – Spot filter

• Default Spot filter criteria have been changed.

Review – Spot normalization

• Interface has undergone slight changes to improve usability and consistency.
• Fitting error in the Normalization table has been made symmetrical to allow selection of both under- and over-expressed spots with high or low Fitting error. The Fitting error of a spot can be defined as the vertical distance of the spot to the Median (red line) in the Normalization plot. Mathematically, for spots above the Median, the Fitting error corresponds to Vol Ratio*Factor. For spots below the Median, the Fitting error is calculated as 1/(Vol Ratio*Factor).
• Correlation and CV in the Normalization factor table were replaced with Fitting and Mean fitting error (MFE), to make evaluation of different normalization options easier.
  Fitting corresponds to the slope of the linear regression between the Normalized spot volume in the sample image and the Normalized spot volume in the reference image, when the fitting line is forced through the origin. Optimal normalization should give Fitting values close to 1.
  MFE is the geometric mean of the Fitting errors (in the Normalization table) of all spots in the particular image. The magnitude of the MFE will depend on the ‘Display ratio as’ option in General options > Quantification. Nevertheless, lower MFE will indicate better fitting. It is therefore recommended to look for minimized MFE when choosing between normalization options.
• For DIGE experiments, internal standard images are now also normalized (to the internal standard image that best represents the mean values of all internal standards). This option can be deactivated in the Workflow options for Spot normalization.
• Ratiometric normalization now only considers the (50%) most conserved spots in all the images of the normalization group. This is also the case for the normalization of the internal standard images in DIGE experiments. For within-gel normalization in DIGE experiments, all spots are taken in consideration.
• Both non-normalized volumes (visible by default) and normalized volumes (not visible by default) can now be displayed in the Normalization table.
• Annotations can now be visualized in the Normalization table.

Results

• Spots in the Anova table are by default sorted according to the p-values in the Anova column (or the Interaction column, for two-factor analyses), from smallest to largest.
• Small improvements in the Filter by values feature.
• Small improvements in the Interaction plots and Expression profiles.
• Column settings have been added to the Expression ratio table.
• Plot display options, and the column visibility and sorting of reports are now stored and restored when navigating between workflow steps or reopening a project.
• Annotations are now conserved after editing of alignment, detection or normalization.
• Spot sets are now conserved after editing of alignment, detection or normalization. The tooltip on the Spot set name (column header) indicates whether the set has been modified since its creation (for sets created from Combine spots sets), or whether the data used for its creation has been modified (for sets created from Filter by values).

Picking

• The matches can now be hidden.
• Spots to pick can now also be defined based on annotated spots.
• Images are automatically unwarped when selecting the Ettan marker tool, to ensure that the Ettan markers are visible.
• When exporting the pick list for the GE Healthcare Ettan picker, a table is created with the coordinates of the Ettan markers.

Display area

• In the Options, it is now possible to display an Overview window that shows where on the gel the visible area (region of interest) is located.
• In 3D view, regardless of Mode, it is now possible to translate the images by pressing the Ctrl key while holding down the scroll wheel and dragging.
• 3D views are now normalized based on the normalization Factors.
• When images are only shown in 3D, features that are not available in the 3D view have been removed from the display options.
• Minor changes in the management of the Modes.
• Measure tool is now available in all workflow steps.
• For each workflow step or sheet (e.g. different analyses in Results), image layout as well as display options for Grid, Profile, Overview, Scrollbar and Spot shape are saved and restored when the user comes back to the sheet.
• It is not possible to display the Profile when “Keep image data in memory” is deactivated, to avoid memory-related issues.

Options

• The Workflow options have been reviewed and are better managed. Some options were removed or renamed to make them more comprehensible.

Performance and usability

• Several changes have been made to optimize speed and memory usage.
• Navigation between workflow steps has been optimized.
• The reactivity of the software during image or overview movements has been improved.
• The software now stores user-customized window sizes and positions. To restore the default layout settings, use Reset all in General options > Display > Layout.
• Keyboard shortcuts have been adapted, simplified and documented (on the product website).
• Minor terminology changes have been made to increase clarity and consistency.
• The same typeface is now used throughout all interface elements.
• For better accessibility, font sizes of the various interface elements now adapt to the Display option chosen in the Windows Control Panel.
• When no image is selected, the Adjust contrast window displays an information message.
• Columns “pI”, “MW” and “pI MW” (for pi/MW calibration) can now be visualized in virtually all reports throughout the workflow.
• Cursor information now also displays the raw pixel intensity in addition to the calibrated pixel intensity.
• Modification date in the Projects interface has been replaced with Opened date.

Fixed bugs

Quality control

• Error in the initialization of the highest pixel value for .gel files. This notably impacted the results for display of saturation in the Quality control step, and subsequent detection and quantification.
• Displayed images could be inconsistent with recently removed/added images.
• Color overlay for saturated and background-clipped pixels onto the images was not properly displayed.
• Small corrections have been made in the feed-back panel at the left.
• In Image edition, it was possible to get two times the Rotate free dialog.
• In Image edition, an empty crop zone could be created on double-click.
• In Image edition, it was not possible to Undo a Change resolution.
• In Image edition, it was possible to reduce the resolution to a value lower than 100 dpi.
• It was possible to quit Image edition without confirmation, while still being in Change resolution mode.
• When changing the resolution of images in an analyzed project, the alignment and detection were not reset.
• When adding, removing or editing images, the image normalization was not reset.

Alignment setup

• Crash when redefining a different reference image, on a data set that was already aligned and detected.
• Crash after choosing Ungroup, on a data set that was already aligned and detected.
• Software could crash when creating a group and then selecting an image with Shift.
• Image order in the Display area did not reflect the order of the images in the tree view.
• Ungroup changed the order of the gels.
• When gels were grouped and pane layout was changed from the default, pane layout was reset when dragging gels to define a new reference for the group.
• User could select items at different levels in the tree view.

Alignment

• Crash when pressing Esc to interrupt the first alignment.
• Crash after Undo of an Unalign action on a project with hierarchical alignment setup.
• Memory leak after Reset modifications of the Unalign action.
• Warped view and match vector positions were not always properly refreshed.
• Display choices were not correctly restored when restarting the software.
• Area of interest was not conserved when going to a particular Alignment from the Results step.
• Alignment counter (Alignment X/Y) didn’t take into consideration if the Intra DIGE align option in the Workflow settings was activated or not.
• Contrast adjustment on the reference image, using the Contrast ruler, was not stored and restored when switching between alignment steps.
• Blink option in Display didn’t work when the previously selected option was Side+Side or Dual+Blink.

Detection

• Software could crash when Warp was activated in Detection, when spots went past the border of the image.
• Spot volume could be wrong when spots contained pixels with intensities lower than the background.
• After matches were edited in the Alignment step subsequent to a first detection, the detection parameters displayed in the Detection step were erroneous.

Review

• In certain conditions, Group table was not updated after edition of spot normalization.

Review – Spot filter

• When restoring the Default filter, there was an inconsistency between the value and type of filter.

Review – Spot edition

• Delete > Delete selected… option did not delete the selected spots.

Review – Spot normalization

• Normalization options were not always properly managed.
• Reference image used for normalization was not always updated when settings were modified in spot detection, spot filtering or spot edition.
• In specific conditions, the selection of spots for Ratiometric normalization could be lost.
• First edition of the markers (Spikes) selected for Spike normalization was not applied to the Normalization plot, Normalization factor table and images.
• Images were not visually aligned, which also resulted in varying parts of the images being displayed in 3D view.
• Several corrections in relation to spot selection.
• Spot colors in 3D view were not updated to reflect the colors in the 2D view and the Normalization plot (and the choice of normalization method).
• Position of the Ratiometric normalization (now Markers) column in the Normalization table could change when switching between different normalization options.
• Crash when displaying a Scatter plot with Volume ratio versus Mean volume, and then changing the option for displaying ratios (General options > Quantification).

Results

• For a two-factor design, when the subject factor was on the top factor in the Experimental Design matrix (i.e. on Factor A, the first created factor), then the ANOVA p values were not correctly calculated in the results. Some values showed 1.#QNAN or 1.#IND values.
• When creating an additional analysis in the Statistics tab, using a design with subjects, it could happen that the images were not opened and the reports were empty.
• Small p values (<10e-8) were not correctly ordered on column sorting.
• For DIGE projects with an internal standard, the values in the Values summary table in Results were not consistent with the selected ‘Display ratio as’ option (in General Options > Quantification).
• For DIGE projects with an internal standard, the interaction plots always displayed Vol Ratio expressed as Log10 values, not considering the selected ‘Display ratio as’ option (in General Options > Quantification).
• Current tab was not correctly restored after opening and closing the Image view.
• 3D views were not correctly redrawn when switching between Analysis tabs (they appeared black).
• Scroll bar of Experimental design view was not re-sized when its window became smaller.

Picking

• Ettan markers were not saved.
• After reopening software in Picking step, images were warped but match vectors were not superimposed.

Display area

• The software could get ‘Out of memory’ with 3D view display after switching between workflow steps (more than 40 switches).
• When exporting the whole image to the clipboard, only a part of the image could be exported, depending on the zoom factor.
• The display options Warp image and Display spots were accessible in Image edition and Alignment setup (where they were not applicable) and could cause crashes when selected.

Image view

• Selection of images used in the Image view was not conserved when closing/opening the software.
• Selection/deselection of images used in the Image view could make some pane headers disappear.
• Crash when opening General options while in the Image view mode.
• Disabled spots were visible in Image view, when Image view was accessed from workflow steps without spots (Quality control, Experimental design, and Alignment setup).

Annotations, spot sets, validation columns

• Crash on import of annotations on spots with existing labels.
• Combination of Validation sets (with ‘Combine spot sets’) gave erroneous results when some spots were excluded from the analysis.
• Spot sets created from ‘Filter by values’ displayed inconsistent filter criteria in their tooltip.
• Spot Sets created from ‘Combine spot sets’ displayed wrong operator in their tooltip.
• When a spot set was edited, and the operation then Undone, the ‘Modified after creation’ status in the spot set’s tooltip was not updated.
• Validation columns displayed numbers instead of usable labels when exported or displayed as labels on the images.
• In Demo mode, ticking a box in an Analysis-ID validation column showed the Demo mode message for each selected row.

Tables and plots

• Print table didn’t work for users having Chrome as the default web browser.
• Numbers on the vertical axes of plots could be truncated when the window size of the plot was changed (vertically).
• In a table without any rows selected, the Ctrl+A shortcut selected all rows, but not the corresponding spots on the images.

General

• On some specific succession of steps, the state of the workflow bar and the effectively active step could become inconsistent.
• Incorrect value displayed for Pixel intensity in Cursor information and Status bar.
• When displaying license information, Copy to clipboard did not work.
• Correction on closing Workflow settings window using keyboard (Enter or Escape).
• The Busy/Waiting cursor could flicker on some actions.
• Small correction on flashing behavior of modal dialogs.