Tracking Objects with the Measurements (Object Intensity)

Hi,

I am trying to make a pipeline to track a few nucleoli in my cells at a time, (there are maybe 20 cells in a frame at a time). They are fibroblasts taken with a 20x objective so the nucleoli are pretty small, but so far I have been able to create a pipeline that will reliably pick them up. The issue is that now I would like to track them. Since I ran a module to enhance dark spots (the nucleoli), it returns an image where the nucleoli in any give nucleus have varying intensities. I would like to use the measurements method within the track objects module to distinguish nearby nucleoli. I added a MeasureObjectIntensity module before the track objects module, but for some reason it will not perform the tracking. It will pop up an original image where it assigns all the nucleoli a number but it will not try to track the second image.

To troubleshoot I tried to simply track the nuclei using CP (we currently use other software to track the nuclei). I was able to easily track the nuclei using the distance and overlap methods but could not get it to go past the first image again with the measurements method (the LAP method will run but I have not gone through all the troubleshooting to get the object assignments to be correct).

Is there something that I’m missing? Is there another module that must be added in order to use the measurements method?

Also, I am running CellProfiler 2.0 from a cluster.

Thanks so much!

-My

Hi My,

Would you mind posting your current pipeline, plus a few sample images so we can give it a try (perhaps 10-20?). You can zip them up as one file if that’s helpful.

Regards,
-Mark

Hi Mark,
Thanks so much for your quick response!!

So I had to include the images in two separate zip files since they are so large. I am including an extra image set (FITC)that is not necessary for the tracking portion, because our pipeline calls for it.
The nuclei images are the CFP ones.

Although, I can capture the nucleoli, they do not look amazing.
For now I just want to get the nuclei tracking to work (I know that getting the nucleoli tracking to work is going to take a lot of tweaking), so that is what is included in the pipeline.

Thank you so much for your help!!!

-My
Segment_NucleoliDetect.cp (22.9 KB)
FITConly.zip (7.13 MB)
CFPonly.zip (6.61 MB)