Outline DAPI nuclei in CellProfiler

Hi, I am new to pretty much everything here. But I think I am trying to do the same thing as you are. Just to clarify are you trying to get the CellProfiler to outline the DAPI nuclei on cell and then use that same area in a different area of the image to quantify the intensity of the stained DNA damage? I am currently trying to measure the intensity of different signals from different antibodies within the nuclei. I have noticed that this program is structured similar to code which is my weakest area. So if anyone could help me out, that would be great and save me from having to quantify over 160 images by hand.
Thank you so much.

Welcome to the forum, @Vickie_Hwang!

Since your question is specifically about CellProfiler, whereas the other thread was about an ImageJ macro, I moved your post to a new topic and tagged it with cellprofiler so that CP experts will be more likely to see it and respond.

Did you make a pipeline that you can share? Or share a sample image and describe (or annotate by hand) the desired segmentation you are trying to achieve?

Yea, that’s why we use code as much as possible. Essentially, what I’m trying to do is measure area within each nuclei, but it would be easy to assess mean grey value, which I believe is integrated intensity based on area (Intensity/area). If I can get the code to work I’ll send it your way, all you would need to change is the Set Measurements line to make it read the appropriate measures you desire. But let me see if I can get this to work.

Steve

Vickie,
You can also get FIJI for free and just look up on youtube how to use. FIJI is produced by NIH. But don’t download ImageJ, do FIJI as it is better than imagej. If you want the code you can have it, sorry I missed the comment about Cellprofiler.

Steve

Immunofluorescence Outlining DAPI.cppipe (36.8 KB) 190712_80min_3a5 atri_1.tif (4.2 MB)

So this is the pipeline that I have created so far. And I have also attached a sample image that I have been trying to quantify. I am trying to outline the blue dapi and then overlay it to the green and red channels. The bottom right corner is just a reference of all three channels overlayed.

I have been using ImageJ before but it just takes a long time to do it by hand, which is why I have been looking for alternate solutions.

I have been able to overlay the nuclei outline on the red corner perfectly fine. But for the green corner, I have been having issues with lining it up because it is in between a pixel and I cannot do half a pixel. The other main issue is that the program is having a hard time identifying the nucleus from the grayscale image. It will either break up the nuclei into many parts thinking it is a clustered object or it won’t recognize it at all.

New Text Document (5).txt (1.7 KB)
We got our code to work. Actually, I believe it worked all along. I just didn’t have enough sense to look at the %Area as opposed to just the absolute area, which I guess is how imagej works. You might have to tweek it some for your project, but this is all the help I can give you. I’m very new to this so I didn’t really have any idea what you were talking about. Sorry I cannot be of more assistance. I hope this works for you!

Steve

Thank you so much for your help!