The use of IndentifySecondary



In my study, some cells express GFP and others have no expression. I use CellProfiler to analysis the intensity of the GFP expression. First, I use the IdentifyPrimAutomatic Display to outline nuclei. Then, I used the IndentifySecondary, but I found the cells that haven’t expression has the nuclei’ outline that shouldn’t have in the picture. How can I do to analysis the cells that only have GFP expression?

Thanks for your help.


Hi Chen Na,

You can use the FilterByObjectMeasurement module to filter nuclei that have GFP.

  1. Define nuclei
  2. Measure intensity in GFP channel on Nuclei objects
  3. FilterByObjectMeasurement of Nuclei that have high GFP
  4. IdentifySecondary on filtered nuclei



Hi Mike,

I don’t know how to operate the two and three that you told me. Shall I use the MeasureObjectIntensity or MeasureImageIntensity to measure intensity? How to measure intensity in GFP channel on Nuclei objects?

Thanks for your help.

Chen Na



You want to use MeasureObjectIntensity to measure the intensity of the nuclei in the GFP channel. Load the GFP channel in your LoadImages module and give it a name, for example OrigGFP. Then, in the MOI module, choose the OrigGFP image and Nuclei as objects. This will measure the GFP channel for each Nuclei. You will need to read the help to undertsand how the FilterByObjectMeasurement module works.