Neuronal Intensity Distribution

Hi, I am looking to use the MeasureObjectIntensityDistribution to map the spatial distributions of neurons. I’ve identified the large object -> total organoid area and have set that as the center. I’ve also identified neurons as the primary object to measure. However I get this error message, ideas?

Hi Samir,

Could you upload an example pipeline and a sample images so that we can try to reproduce this error? It would also be helpful to know which version of CellProfiler you’re using.

Thanks!

Hi Pearl

I’m using version 4.0.4. I have not properly set up MeasureObjectNeighbors or the Skeleton modules but they should be separate from this.

Thanks!

Neurites.cppipe (34.7 KB)

Hi @Samir_Gouin,

I think that the error you’re seeing is from a bug in CP 4.0.4 that affected visualizing results from modules like MeasureObjectIntensityDistribution. I ran your pipeline using an unrelated image in CP 4.0.6 without any errors from MeasureObjectIntensityDistribution.

If you can, I’d recommend updating to CellProfiler 4.0.6. If you can’t do that, try turning off the window (click on the eye next to the module). The measurement should proceed normally as long as you don’t ask CellProfiler to show the window.

Does that work?
Best,
Pearl

Hi Pearl,

That helped, thanks! I no longer get the error message. My results are a bit weird (attached image). It appears it is not creating a heat map that spans the entire organoid area, rather, just a few nuclei in the center. In my MeasureObjectIntensityDistribution settings, I’ve selected the total Organoid Area to be used as the center so Cell Profiler should use the center point of the total Organoid Area. Any ideas? Maybe the issue is in the IdentifyPrimaryObjects for the Organoid area?
Thanks,
Samir

2019.09.12 cerebral 2mo stain2 A.08_2019.09.12 cerebral 2mo stain2 A.08_2019_10_05 Batch 2019_05_27 B373 B374 Ast 234KO 100d MAP2_488 S600_546 Syn_647 Hoechst n7 Mergeddistribution.tiff (7.7 MB)

Hi @Samir_Gouin,

If I’m understanding your neurites.cppipe correctly, it looks like you’ve selected Nuclei as your primary object to measure the intensity distribution of the neuronal channel relative to the center of the Organoid object. From your initial question, I think you want to measure the distribution of the neuron channel within the Organoid object in order to get a heat map for that entire region. You may find the help module for MeasureObjectIntensityDistribution helpful to see different examples of how this module can be applied.

If I’ve mis-interpreted your question, then a bit more explanation of your biological problem and what you’re trying to measure along with an example set of image inputs would be helpful to troubleshoot the pipeline. Good luck!

Pearl

Hi Pearl,

Yes, I want to measure the distribution of nuclei within the organoid area. Does this look like the right approach? I was not able to find a specific example and receive an error message when I try this.

Hi @Samir_Gouin,

Since you’re using the Organoid_Area_identify as your object to measure the distribution of the fluorescence of the neurite channel, you can:

  • select the appropriate image using the checkbox for your nuclei channel
  • then assign objects to measure as Organoid_Area_identify and use the same object as the center.

This approach works for me using mock data to replicate what you’re describing. If that’s not what you’re looking for, you may want to submit an example pipeline and sample images or sign up for one of our Virtual Office Hours in the next few weeks. Good luck!