This is Fiji software tutorial for SpatialTime package.
- Open Fiji software. You should see a similar interface:

- Import your sample “tissue_hires_image.png” image into Fiji by going to File > Open. Pressing Ctrl+O or clicking and dragging the image into Fiji interface also works. You will see a similar window:

- Go to Image > Color > Stack to RGB. Another window might pop up. Then, the reference line is ready to be drawn. First, double click in the colored squared in fiji interface.

- A window with multiple colors will show up. In this example we’ll use red. Double clicks in one of the double-colored squares in inferior section. Then, make sure to set the foreground Color equals 255 while other are 0.

- Select the Pencil Tool in Fiji interface and drawn your reference line.

- Press “Freehand selections” in Fiji interface:

- With freehand pencil contour around your reference line. That way you will export mainly the image XY coordinates associated with your reference.