Thanks for your quick reply, Chris. I really appreciate your help.
So my raw images really are 8-bit, but I even redownloaded my image off of here and got the same results. I’m also trying this on a second computer with fresh installs now. Maybe there is some other setting referenced by the script I am missing? Also, I’m not sure what you mean by setting a range in the Brightness and Contrast tool. This is the window that opens with Shift+C, yes? But seems like that is just an issue for 8-bit conversion.
Below are images processed using the file downloaded from my original post. Here is what I get running the Apply_DOG_Filtering.py script with sigma values of 10 and 5. Something seems to be wrong with the upper threshold:
Here is what I get using the GDSC DoG plug-in with and without the contrast enhanced:
When I submit the second one to ridge analysis using your parameters it seems to be working, but I end up with different diameters:
Maybe this is because I have a scale set on my images? However, to my eye, it looks like I’m losing some of the fiber ridge definition at the DoG step.
What do you think?