Measure Neurons module and Morphological Skeleton

Hi there,
Since I am new with image analysis and Cell profiler I would appreciate a bit of help.
In the three images that I have uploaded I have three different states of neurons after treatment. My imaging is not neurite targeted (simple alive cell staining) but the difference by eye is obvious I just can not put it to numbers.
I would like to measure the Neuron body numbers and the total axon length and the branches. I went through all the instructions and the forum with the related topics but I can not solve my problem.
I am using Cell Profiler v4. but I can not see the measure neuron module, I know that I have to do Morph and or Morphological Skeleton but I do not obtain any result either with grayscale images or with binary mask. Although in the pipeline that I have uploaded by Identify Secondary Objects I can detect the axons in grayscale images
I guess I am missing something fundamental.

Thank you in advance

B02_T0001F005L01A01Z01C01.tif (1.9 MB) B03_T0001F005L01A01Z01C01.tif (1.9 MB) B08_T0001F005L01A01Z01C01.tif (1.9 MB) Neuritestest3.cppipe (10.7 KB)

Hi @sadikoglueldem,

I checked your pipeline & images. To get your Morphological skeleton you need a binary image & it can be obtained by “Threshold” module. So before your Morphological module use this & it gives you the skeleton as shown in the screenshot. later the skeleton can be measured using the MeasureImage Skeleton. PFA modified pipeline.


Neuritestest3_lb.cpproj (468.0 KB)

Hope this helps.

Regards,

Lakshmi
www.wakoautomation.com

Dear Lakshmi Balasubramanian,

Thank you very much fro your reply. Your pipeline worked good for me finally I just had to change the threshold strategy from Global to Adaptive and Sauvola to include some fine neurites that were undetected. What I have noticed now is that I have a lot of granules type structure on my axons which are not selected in primary object (magenta circles) in file GranulesCells but are included in Threshold output and the subsequent steps of MeasureImageSkeleton. Could you please suggest away of removing them from the MeasureImageSkeleton.

Thank you very much

Hi @sadikoglueldem,

Great!!
You can try suppress the speckles using “EnhanceSuppressFeatures” before thresholding or get rid of during the thresholding based on the size.

Regards,

Lakshmi
www.wakoautomation.com