Analyzing perinuclear clustering of mitochondria

Hello. I am a new (and completely inexperienced) user of cell profiler. I learned about the software from a publication by Cataldo et al, 2010 and I have to say that it sounds like a really helpful tool. I am interested on using the software to measure mitochondrial distribution around the nucleus, like they did in the publication. After I did some research, I found out that the pipeline they used in this publication has been uploaded in the forum but I have to admit that I encountered some difficulties while I was trying to analyze my images, since I am getting an error in the beginning of each analysis (see attached image). Therefore, I was wondering whether it is possible to get some step by step help on the issue. If someone can help me I will really appreciate it.

Thank you in advance for your time.

Hello!

Do you have any images loaded via the “Images” and “NamesAndTypes” Module? The latter has a “Update” button, if you get an error or do not see your image set table, you need to recheck the settings for defining the image sets.

If this is not the issue, uploading your pipeline and some sample images is always useful :slightly_smiling:

Hi @Fabba123. Thank you for your response. Let me guide you through what I have done until now so we are on the same page. First I downloaded the pipeline Cataldo_2010.cp (18.1 KB) that was uploaded by @David_Logan and I started working on that. After I removed the “ExportToDatabase” module I was still getting an error (see image below).

After I spent some time trying to figure it out, I realized that I was getting that error due to one of my settings on the “NamesAndTypes” module.

After I changed the “Image set matching method” to “order” I was able to press update and see the a table.

After these adjustments I was able to press “Analyze Images”. The result I got can be seen below.

Nevertheless, I am not sure what I am looking at, since there was no image associated with that analysis. Is there a way to export an image showing the concentric circles like Cataldo et al, 2010 did in figure 3B of their publication? Also, is there a way to set the software to increase the diameter of each cycle by 1 μm (for example) from the previous one?

I am also uploading the images I used for this test analysis and the pipeline. The images were previously processed with ImageJ for Sharp, Find Edges and Make Binary, as Cataldo et al, 2010 suggested.

Actin

DNA

Mito

Pipeline
mito_distr.cppipe (19.3 KB)

I will try and help far as I can (general aspects of CP - but I have no experience with mitochondria or this pipeline)

  1. You only have one image set loaded. The results displayed in the window you posted pertain to THIS image set.
    You can see what Image was measured (MitoBinaryMaskNuclei AND Mito_orig), the objects measured (cells), the list of bins (1-8 in each image, with “bin count” as the total number of bins).
    “Fraction, Intensity, and COV” are your actual measurement outputs - but I have no experience with this module.

  2. This is only one way to view CP results. You will also find all measurements in the resulting spreadsheet files (.csv). I would strongly recommend also using the ExportToDatabase module (Settings: use SQLite and make sure to create a properties file!), as it will allow easier viewing of large data with CellProfiler Analyst, if you like to at a later time.

  3. While it is possible to overlay a LOT of information onto images with CP, I think the image in the publication you are referring to was not made with CP.

That’s all I know, the Pro’s will be able to help you out more :smiley:

If you use the Beta version here then you see a visualization from MeasureObjectRadialDistribution (which we renamed MeasureObjectIntensityDistribution recently) as long as you choose “Add another heatmap display”. In the release version (version 20140723) of CellProfiler there is no visualization of this.

Unfortunately I am not able to open the beta version of CellProfiler. I am getting the following error.