WebAlign

WebAlign is used to register brain sections to a reference brain atlas using a linear registration method. This involves two key steps:

1. First, establish the series cutting angle using key anatomical landmarks. The angle is usually stable across the series unless sectioning is performed in two or more blocks. Once the angle is established, adjust the plane through the atlas to match this angle for all the sections.

2. Second, manually register each section to the atlas by etablishing its position and then adjusting the size and position of the atlas overlay to match the underlying section.

Once the atlas is correctly registered to the sections using WebAlign, WebWarp can be used to make in-plane nonlinear refinements to match anatomical landmarks.

Data from: Laja et al. (2025). Distribution of parvalbumin-positive interneurons in the normal adult rat brain (v2) [Data set]. EBRAINS. https://doi.org/10.25493/MB6S-K0S

How to use WebAlign?

1. In your project, Open WebAlign by selecting “Continue to WebAlign”.

2. Select the atlas orientation that best fits your sections using the drop-down menu in the Control Panel (coronal, sagittal or horizontal). This adjusts the plane through the atlas template for all the sections.

3. Select a section to register in the filmstrip. Start with the first and last sections in the series (see tip below). The Main Window shows the selected image with an atlas overlay. The colour of the image border reflects the registration status (white - not registered, pink - in progress, green - registered).

Tip

It is recommended to register one section in the beginning of the series and one section in the end of the series as the first step in the registration process. This is because WebAlign automatically updates the position of remaining sections based on the section numbering, making the registration process more efficient.

4. To register a section, move the plane through the atlas to the section’s approximate position using the yellow dots in the sagittal, horizontal and coronal windows (Navigation panel). The main window should now show an atlas overlay that roughly matches the appearance of your section.

5. The main window supports mouse drag in multiple modes for adjusting the atlas overlay to match the outer borders as well as anatomical landmarks within the section (both translation and transformation are supported).

• Markers are positioned using the “Space bar”. Markers are crosses initially and are the anchor point of transformations to the atlas overlay. The “Esc key” can be used to remove the marker (in full screen mode the “Esc key” escapes full screen).

• If there is no marker, or the marker is a cross, mouse drag slides the cut in its plane (translation).

• With a cross in place, press the Up and Down arrows or Left and Right arrows to activate stretch mode (a line will appear) or choose direction with the arrows present in the top left corner of the mian window. Adjust the atlas overlay to match your section. The panel can be resized towards the left (common border with Control Panel) and towards the bottom (common border with Filmstrip).

6. Once a section is registered to a satisfactory standard, tick the box in the filmstrip to save the registration. This will change the registration status from pink (in progress) to green (registered). This automatically propagates the registration to the remaining sections to help with scaling.

7. To establish the series cutting angle, look through sections with key anatomical landmarks. Once the angle is established, apply this to all the registered sections. The angles will automatically propagate to remaining sections.

8. Go through all the sections, scaling and refining the positions of the atlas overlays, and ticking the boxes on the filmstrip to save the registrations. Registration is complete when all the images have a green outer border.

Note

When jumping from one section to the other, wait a few seconds for the image to load.

Note

The “undo” button allows you to go back to the saved position if necessary.

7. To complete the registration, apply fine adjustments with WebWarp. This step is optional.

Short keys

To do this	Press	Description
Place marker	Space bar	Markers are the anchor points of most transformations (stretch and rotate)
Remove marker	Esc	Removes a previously placed marker
Horizontal stretch from maker	Left/Right arrow keys	Marker becomes a vertical line, mouse drag horizontally resizes the cut
Vertical stretch from maker	Up/Down arrow keys	Marker becomes a horizontal line, mouse drag vertically resizes the cut
Rotate around marker	PgUp/PgDown	Marker becomes a cross with a surrounding arc, mouse drag rotates the cut
In plane adjust	Click + drag	If there is no marker, or the marker is a cross, mouse drag slides the cut in its plane (translation)

Control panel

Button	Function
Pan/Rotate step	Allow to change step size for pan or rotate functions
Overlay Slider	Opacity of the atlas overlay, when fully opaque, it becomes an outline
Overlay color	The outline color
Filmstrip slider and color	The above settings, applied to the filmstrip
Segmentation/Template	Allow to switch between visualisation of the atlas segmentation or the atlas template
Undo	Undo the current alignment to the last stored position
Redo	Return to last previous position
Clear	Reset the current alignment to the default position
Center	Adjust and center the cutting plane through the atlas to match the orientation selected in the drop-down menu (coronal, horizontal or sagittal)
Angles	Switch between Coronal/Sagittal and Horizontal planes and displays angle values. Angles are color coded to their respective planes.
Apply to all	applies chosen angles to all sections in the series
Diagrams	Open a new window with diagrams displaying the relative position of the sections in atlas space, section spacing, primary and secondary angle.

Note

The border of the control panel can be dragged horizontally, allowing resizing of the panel and the main view.

Filmstrip

Drag horizontally to see series, click on a section in order to load it into the main view. Save your registration by clicking on the small square on top left. The top border of the filmstrip can be dragged vertically, allowing to resize the panel and the main view

Navigation panel

Shows the three standard planes centered around the midpoint of the current alignment visible in the main view.

The rectangle of the current cut is projected on each standard plane as a yellow line/rectangle/parallelogram. A small yellow circle represents the midpoint of the projection.

Drag the midpoint around to move the cut.

Drag anywhere else to rotate the cut (inside the given standard plane, around the midpoint)

Diagrams

Press the “diagram” button to display graphs showing the relative positions of the sections in atlas space, the spacing of the sections relative to each other, and the primary and secondary angles applied to the sections. These can be used as a guide in the anchoring progress to ensure sections are registered in the correct order, that they do not overlap each other in atlas space, that the angles are consistently applied to all the sections.

More details can be found here: https://webalign.readthedocs.io/en/latest/