Analyzing Spheroid Images

Hi All,

I’m working on a pipeline for quantifying cell migration in spheroid images. I started by simply using the MeasureImageAreaOccupied module and I was able to get results from it, but I realized that the actual area of the spheroids is not the most reliable metric for what I’m hoping to capture. What I really want to measure is the degree to which individual cells migrate away from the central spheroid (I have a few images at the end of this post that show what I mean by this a little more clearly). I have been thinking of either trying to quantify the degree to which the edge of the spheroid is round compared to a less uniformly curved border, or if possible I would like to be able to identify cells outside of the spheroid and get a distance from the spheroid, I just don’t know how/if I can do this with CellProfiler. If you know of any ways to accomplish either of these, or a better way to quantify this, I would really appreciate any help!

In both images the area of the spheroid is pretty similar, but you can see that the second image has many cells that have visibly moved outside the bounds of the spheroid, while this is not the case for the first image.

I’m not sure that I’ve explained this super well, so if anything is unclear or just doesn’t make sense let me know and I can clarify. Thanks!

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If I understand your question correctly, you may want to try:

  • IdentifyPrimaryObject for the spheroid (use some limits of diameters to ignore small objects)
  • IdentifyPrimaryObject (again) for the small cells (use some limits of diameters to ignore big objects)
  • RelateObjects, set parent object as the identified Spheroid, child object as the identified small cells. Check the option for calculating parent-child distances.

Then export the data with this relationship distance and see if this metric reflects what you aim for.

Hope that helps.

That looks like it should work perfectly, thank you so much!

Hi @NickSeyler,

We’ve been working on this type of thing looking at invasion of cells from a spheroid into the surrounding matrix using Lightsheet. You may find some of the methods in this preprint of interest:

We’re just handling reviewers’ comments but feel free to post any questions here or get in touch off list.


This could be really helpful, I’ll definitely look into it. Thanks!