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How to Subtract two images created from Oval ROIs?

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I am trying to subtract two images that have each been created from a user-selected oval shaped ROI. I can preform the image subtraction using the original images, and then selecting an ROI (I can only get rectangular ROIs to work) from the resulting subtracted image and converting that ROI to an image. I'll attach an example program and some sample images for clarity. Any suggestions would be greatly appreciated!!


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