Nuclear/cytoplasmic intensity




I am looking to analyse staining intensity for nuclear alone and cytoplasmic alone staining on one channel (red). I have a nuclear (blue) and cytoplasmic (green) stain to identify the objects and analyse the nuclear intensities of the red stain and this works fine, however I am trying to analyse the cytoplasmic intensity without including the area that is nuclear, is there a way to exclude this from the cytoplasmic intensity of the stain of interest in red using the outlines identified from identifying the nuclei in blue, cytoplasm in green?

Thanks in advance


Hi Charlie-
If your pipeline is set up using IDPrimary to find nuclei then IDScondary to find cells (which we recommend), then IDTertiary will give you the cytoplasmic areas; if not, you can use MaskObjects to exclude nuclear area from cell area but know that the correspondence between cells and nuclei won’t be 1:1.
Good luck!


Hi Beth,

Thanks for the response. What do you mean by the correspondence between
cells and nuclei won’t be 1:1?



If you find the nuclei and the cells in two separate IDPrimary steps, the boundaries of nuclei and cells will be totally separately determined- one nuclei may have two cells associated with it, or a cell may have two nuclei, etc. If you use IDPrimary then IDSecondary, it will find one and only one cell per nucleus (aka a 1:1 correspondence).


Ok I get you now. I have it as primary then secondary for identification,
so that should be fine. Thanks!


Here is a pipeline I have used to do a similar thing. Hope it helps.

ratio-nuclei-count-pipeline-mcclean.cpproj (174.6 KB)