How to handle differences in magnification

[Ping-lin14]

Hi Chandler,

Thank you very much for developing VALIS. I have a question I’d like to ask.

I am working on registering H&E and IHC images, but there is a large difference in magnification. The H&E image is at 10×, while the IHC image is at 40× (the IHC corresponds to an ROI within the H&E image). In this situation, what would you recommend?

Using the standard workflow, I tried a combination of H&E at 20× and IHC at 40× with micro-rigid registration, and the results were good. However, when using H&E at 10× and IHC at 40×, the difference seems too large for successful registration.

My current approach is to manually crop the H&E image using GIMP before performing the registration.

Best.

[cdgatenbee]

Hi @Ping-lin14,

It's a little counter-intuitive, but I've also found that sometimes using the lower resolution images can yield better alignments. I believe this is because the rigid registration ends up focusing on prominent global features, resulting in better alignment that gets refined in the following non-rigid registration (which is done at a higher resolution).

I do have a few thoughts on aligning images of different magnification though, which might be useful:

• The aligned images will end up at whatever resolution your reference image is (either the one you specify, or the one that valis picks). So, depending on what you are using the H&E for, you may or may not want to make it the reference. For example, I was aligning 40x images to an annotated 10x (maybe less) H&E image, but I needed to apply the annotations to the 40x image, so I made the PanCK image the reference so that I could process the aligned IHC at full resolution. It could be done differently using warp_from_to functions, but this way was a cleaner in my case.
• Aligning to an ROI does make this more challenging, since rigid registration may not "see" some of the tissue level features. But, I wonder if it would be possible to create a custom 'ImageProcesser' that can crop the H&E for you. It's hard to know for sure since I don't know much about how the ROI was picked.

Hopefully some of that is useful info, but it sounds like your approach is a solid one. If you have any other questions, please don't hesitate to ask.

Best,
-Chandler