Rolling Ball Background Subtraction, Multiple Channel Issue

The rolling ball background subtraction is spot on for the script I am writing, but I have encountered an issue when using it on multiple channel images. The preview looks like it should, removing quite a bit of background noise, and it even looks fine after I run it. But when I switch to another channel a huge amount of signal is removed from the images.

How can I avoid this 2nd background removal?

I used a 30 pixel ball. And I clicked Yes to the “Do this to all channels” prompt.

Thanks for any advise.


Initial state:

After background removal:

After switching channels:


I am not exactly sure what the problem is… You want to remove the background in all channels - yes? You only see a change in your result after moving from one channel to another - correct?

I would guess - and this is really just a guess - that perhaps it is just a brightness/contrast & display issue and that your pixel levels haven’t actually changed at all. Did you check the histogram and/or adjust again the brightness/contrast?

If you want - just post your original dataset… and provide the exact steps of what you did - and we can try to recreate your issue.



Hey, sorry for the late reply. I checked now and your guess is spot on, it just changes the brightness. So I guess the problem is solved, thank you. But can I avoid that it changes the brightness somehow? I can post the dataset when I get back to the lab next week :slight_smile: