Regions of interest - ROIs

Draw Box

Lets you draw a box ROI. You click in one spot, and drag to another to define the region of the box. By default, the box has a depth of 1, if drawn on a 3D image.

Draw Freehand

Lets you draw an outline ROI. You click in one spot, and drag to add points to the outline. Releasing the mouse will complete the outline.

Magic Wand

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Magic Wand Settings

The magic wand tool lets you create outline ROIs. You click on a spot, and the Magic Wand selects points that are connected within a threshold range.

Options for the Magic Wand tool are provided below the row of drawing tools when the Magic Wand is selected (see above).

In Main View (2D), expand the Thresholding Mode dropdown menu to choose one of the following modes:

1. Bidirectional mode accepts connected pixels within the sensitivity limit above and below the intensity at the selected location.

2. Peak Finding mode accepts connected pixels within the sensitivity limit above the intensity at the selected location.

3. Valley Finding mode accepts connected pixels within the sensitivity limit below the intensity at the selected location.

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_{Selected location (small green square), search limits (large green square), and region preview (blue) for the Magic Wand}

Select a location on the image for the Magic Wand to search from. A preview of the region to be painted is shown in blue (see right). Adjust the Sensitivity slider (or use hotkeys W and R) to change the tolerance for intensity differences. The narrowest range of accepted intensity levels is used when the slider is all the way to the left. Adjust the Search Range slider (or use Ctrl + Shift + Mouse Scroll over the Image Panel) to increase or decrease the area over which the Magic Wand will search. No pixels outside of the search region are included in the painted region. The search range is indicated by the green box around the cursor when the Magic Wand is used (see right). Finally, click on Paint (or press E) to paint the region and assign it to the selected class.

Auto ROI Detection

Auto ROI Detection tool in Aivia allows you to auto-detect regions of interest for analysis. The algorithm identified Regions of Interest (ROIs) by analyzing areas of high variance within the input image. To use this tool, switch to the Auto ROI option located in the row of drawing tools within the ROI Editor.

Options for the AUTO ROI Detection tool are provided below the row of drawing tools when the Auto ROI tool is selected.

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Parameter Settings

To apply Auto ROI detection to the image, set the parameters given below. The default settings should be a good starting point. (Kernel Size=5, Sensitivity =7)

1. Input Channel(s): Select the channel(s) for AutoROI detection that display signals from the majority of cells. For instance, DAPI is often a good first choice as it typically labels most cells. Starting with Aivia 14, users can choose multiple channels to enhance ROI detection. Be aware that the number of channels you can select depends on your computer's RAM. Selecting an excessive number of channels will trigger a warning message about potential system slowdown or crashes.

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2. Kernel Size: Kernel size defines the size of the variance filter applied to the input image. Increase the kernel size for a smoother outline and decrease the kernel size for more details.

3. Sensitivity: This setting adjusts the variance threshold to detect smaller ROIs more sensitively. The table below illustrates the relationship between sensitivity and variance threshold. It's important to note that in a 16-bit image, the intensity range is wider, consequently leading to larger variance thresholds. Users should be able to get similar results using the same sensitivity if they convert images between different bit-depths in Aivia. When using multiple channels, Aivia uses the highest variance value for thresholding.

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Using the Auto ROI Detection

After the right parameters are chosen, click on the Detect button on the right side of the view. The regions of interest from the auto-detection can be seen in the Regions of Interest table.

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In general, the ROI naming order is determined by sorting from the top to the bottom row and left to right column based on their center of mass. ROIs are considered in the same row if they fall within our estimated row spacing.

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Common issues

1. Aivia detects no ROIs: Try to reduce sensitivity. If you already use the smallest sensitivity, that means the image’s foreground and background variances are very close to each other. Try to use another channel if available.

2. Aivia detects the whole image as one ROI: Increase sensitivity to enable the algorithm to effectively filter out background pixels.

3. ROI is smaller than the actual sample size: To address this, one way is reducing the sensitivity. Another way is increasing kernel size to get ROIs with smoother outlines. This is particularly recommended when the marker is smaller than that of the entire cells.

Crop

Creates an image from the ROI by cropping the current image to the bounds of the ROI, and only including pixels that are within the ROI.

Revert

Reverts the position of the ROI back to when it was originally loaded/added. Only applicable to Box ROIs.

Clip to Box

Only available in the 3D view and for box ROIs. Moves the clipping planes to be around the current box, allowing you to remove any channel data or mesh data not inside the Box ROI.

Remove ROI

Removes the ROI from the image. This action is undoable (ctrl+u).

Show/Hide ROIs

Shows/Hides all ROIs

Add ROI

Opens a dropdown with options for adding ROIs.

Store

Stores the current recipe settings in the ROI (in the Recipe Console Regions Of Interest Table only)

Restore

Loads the recipe settings that are stored in the ROI into the Recipe Console (in the Recipe Console Regions Of Interest Table only)

Go to ROI in treeview

Goes to the ROI editor and finds the ROI in the treeview, selecting it. It will expand any necessary parent ROIs, and collapse any unnecessary ROIs.

Add a new box ROI

Creates a new box image in the current image.

Load a mesh as a ROI

Loads a mesh from file to be a ROI. The loaded mesh surface must intersect the image to be successfully converted to a ROI.

Create ROIs from Selected Meshes

Creates a ROI from any selected mesh objects.

Create ROIs from Selected Outlines

Creates a ROI from any selected outline objects.

Start drawing rectangular ROIs

Goes into the 2D Box ROI Drawing mode. If not currently in the ROI editor and/or the ROI editor is not visible, it will be selected and shown.

Start drawing outline ROIs

Goes into Outline ROI Drawing mode. If not currently in the ROI editor and/or the ROI editor is not visible, it will be selected and shown.

Start drawing ROIs with the Magic Wand

Goes into Magic Wand Drawing mode. If not currently in the ROI editor and/or the ROI editor is not visible, it will be selected and shown.

Box ROI

A simple box shaped ROI, that has a width, height, and in 3D, depth.

Outline ROI

A 2D ROI made up of points.

Mesh Roi

A 3D ROI created from a mesh.