Drift Correction with AMT and ImageJ
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Drift Correction with AMT and ImageJ
In cases where there is linear image drift due to temperature problems in the room or stage stability, the following steps may allow you to acquire and process images to remove the drift. If the drift is the result of section warping, then these methods will likely not work.
1. Focus and stigmate as usual.
2. In the AMT control program choose Special Acqusitions/AVI Movie Controls/Setup Movie....
3. Setup is a three step process all in the same window. First click Create AVI File. This will prompt you to select a location for the file and a name. Please save to the largest disk available (not C:/ unless that is all that is available). Click the Set button.
4. In the Limits pane, set the time or size options. Ten seconds is fine for drift correction. Click Set.
5. In the Finalize pane, click Fastest Record No Live Display to assure the smoothest acquisition. Click Set.
6. Follow the instructions displayed: To start saving AVI Press F7 or Use Start Button while a Live Image is being displayed. To stop recording Press F8. If you do not stop the recording it will finish when it reaches the Limit set above.
7. Transfer your AVI stack to your processing computer and open it in ImageJ.
8. Open the AVI file using File/Import/AVI.... Choose to convert to 8-bit. You might want to open and inspect the file to see which frames are best. For drift and noise correction you should only need about 4-8 frames. Close and re-open specifying the frame range.
9. Run the StackReg plugin, which requires the TurboReg plugin (see ImageJ web site for details). This will align the slices of the stack.
10. Choose Image/Stack/Z Project.... Try different setting of the Z-projection to get the result desired. The Standard Deviation Z-projection option will give an image of the standard deviation of the image. It will highlight areas where there is still some variation in the image. In the case of samples like Carbon nanotubes this may be due to Z-axis drift of the nanotubes in the image sequence.
Note that it may be enough to use a single slice from the AVI stack, if drift is minor and the image noise level is low enough.