IdentifyObjectsManually - apply objects over entire stack frame

cellprofiler
stack
identifyobjectmanual

#1

Hi There,

Really loving CP by the way!

I’m working with tiff stacks of fluorescent adherent cells and I’m wanting to identify areas in my image that contain no objects of interest, as background. I want to record the intensity of that background, rather than automatically subtract it (as in the Threshold module). For this purpose, I had hoped to use IdentifyObjectsManually, to simply select background areas myself. However unfortunately there appears to be no option in that module to apply the selected areas across multiple frames.

In fact, in the latest version of the module (3.0) there doesn’t even appear to be a setting to save those identified regions, as objects. At least if I could locate this setting, then I could create two separate pipelines, in the first of which I manually identify areas of interest and load those areas into the second pipeline as ‘ROI’.

Any ideas on how I could solve this problem?

Also open to ideas on how I could automatically identify background regions - I already have a pipeline that automatically identifies cells and creates an object mask of them, so perhaps there is a way in which I can simply define everything outside that mask also as a separate named object?

Many many thanks,
and apologies for the huge rambly question!

(In case it’s useful, here’s a link to a folder with my pipelines and files.
https://drive.google.com/open?id=15R4WUlC_8SovU41FqD_cLEd1kzx78HJJ
Pipeline 1 does cell ID, to produce img122.ROI. Pipeline 2 applies img122.ROI to the video 124, measuring intensity values across each stack…)

Ewan


#2

Hi Ewan,

If you’re already identifying cells, you could use MaskImage in “object” mode (and invert the mask) to generate a background-only image, then MeasureImageIntensity on the masked image- I’d personally then choose the MedianIntensity as my background measurement but other things could be reasonable depending on your system.

FWIW, you can indeed still save object masks in 3.0 using the ConvertObjectsToImage module.