Thanks! And ideally what you described first is exactly how we would want to proceed, but I don’t think it will give accurate cell-cell information due to problems with the slide scanning (stitching artifacts), slight stretching and sliding of parts of the tissue, etc. If you have been able to get single cell accuracy on overlays using downsampled alignment, that is great news, and hopefully we will see similar!
With practice much of those pre-analysis problems will be mitigated, but who knows. The plan is to give downsampling a try first anyway and seeing how the results look due to there not being a quick and easy way of accomplishing the high res alignment that would be preferable.
The objective is multiplex cell by cell quantification of whole slide images using strip and restain methods. That will require very precise alignment of not so perfect images, though the overall tissue structure shouuuuuuuld be the same. Though it won’t be
I have seen this work and accomplished it in Visiopharm, which I have access to, but Visiopharm doesn’t play nice with outputting the aligned images for analysis in other software which makes that usefulness… limited. Plus, figuring out an entirely open source pipeline has it’s own benefits.