If you have an imported FIB-SEM stack which requires manual pre-alignment before using the automatic z-alignment, you can use the image processing function Coarse Z-Stack Alignment for a manual alignment of your stack. See the following instruction.

1. You have opened your (imported) z-stack that needs alignment in ZEN.

1. On the Processing tab, select the image processing function Coarse Z-Stack Alignment.
2. Click on the Setup button.
3. The Coarse Z-Stack Alignment Setup opens.
4. The current z-plane (selected by the Z-Position slider in the Dimensions tab) is displayed in cyan and the following z-plane is displayed in red.
3. If you have an image with multiple channels, select the channel which should be displayed in the Image View with the Channels tool.
4. With the Z-Position control in the Dimensions tab, go to the z-plane where the image needs an alignment.
5. Select the Speed with which your alignment should be performed.
6. Shift the following z-planes with the arrow buttons in the top left of the setup until the x/ y shift in the z-stack seems to be eliminated. Alternatively, you can also use the arrow keys on your keyboard.
7. If necessary, adjust the speed step size during the alignment.
8. Repeat the steps 4 to 6 until all shifts in the z-stack are corrected.
9. Every shift is displayed in the Shift List on the left side of the setup.
9. Click on Finish to save the changes and close the setup.
10. The Shift List with all the shifts is displayed in the Parameters tool.
10. On the top of the Processing tab, click on Apply.

1. You have now aligned the planes in your z-stack manually to correct a shift in x and/or y within the z-stack.

See also

• Aligning Z-Planes Automatically (Based on a ROI)