GDSC ImageJ Toolsets

The ImageJ toolbar contains buttons that active tools or run plugins. A toolset is a list of buttons that can be shown on the toolbar. Toolsets can be changed using the "More Tools" menu button (the last button on the toolbar labelled >>).

Spot Distance Toolset

This toolset contains tools for identifying and measuring spots within an image, for example labelled proteins within a cell. The plugins are part of the GDSC ImageJ plugins package.

Align Stacks Allows a stack to be aligned across timepoints to correct for stage drift. This simplifies analysis of all frames using a region of interest (ROI) marked on a key frame.
Cell Outliner Find cell edges using marked seed points. Best suited to identify round cells with edge pixels either lighter or darker than the cell.
Difference of Gaussians A Difference of Gaussians filter enhances contrast of image features that change intensity over a range of distances. Set sigma 1 to set the upper limit on feature size, sigma 2 to set the lower limit.

Use this plugin to preview the Difference of Gaussians filter on the image to see if spots are visible. The enhance contrast option will rescale the pixel intensities so results can be viewed.

Remember to cancel the plugin otherwise the preview image will be final.
Spot Distance Detects spots within a mask/ROI region. The mask can define multiple regions using a different pixel value for each. Outputs a table of spot positions and a summary table of the number of spots with min/max/average distances.

Spots are detected using the maxima within a Difference of Gaussians image. The Smoothing parameter sets sigma 2; the Feature Size parameter sets sigma 2. Contrast changes (edges) much larger than the Feature Size will be removed from the image. (Setting the Feature Size to zero to disable the Difference of Guassians; spots will be detected using peaks on the smoothed image.) Optionally spots can be removed if they fail a circularity threshold.

Provides options to process the current frame or all frames in the stack. The last identified spots can be shown on the image as an overlay. The region counter is incremented for each run of the plugin allowing regions to be separted in the results
Redo Spot Distance Runs the Spot Distance plugin using the previous parameters (no dialog is shown). The region counter will be updated.
Undo Spot Distance Removes from the results tables the most recent Spot Distance analysis. This allows a bad result to be erased if the user decides the analysis failed.

The Spot Distance toolset is designed to be used interactively:

  • The user can align multi-frame images using the 'Align Stacks' plugin
  • The user should draw a region of interest on their image using the Freehand selections tool
  • Alternatively the user can select centres of cells using the multi-point selection tool and run the 'Cell Outliner' plugin to generate a mask image of the cells
  • The Difference of Gaussians (DG) plugin can be run to adjust parameters for the filter using the preview with 'Enhance contrast' selected.

    Once the radii are chosen the preview can be cancelled
  • The Spot Distance plugin can be run with the chosen DG filter parameters. Selecting the option to show an overlay on the stack allows the spots to be reviewed
  • Further analysis can be performed quickly for each new selection using the Redo Spot Distance button
  • Bad results can be removed using the Undo Spot Distance button

Download the toolset. Toolsets should be placed in the ImageJ/macros/toolsets directory.

For further details on GDSC ImageJ plugins please contact Alex Herbert.