I am quite new to cell profiler. I work with monocyte derived macrophages and I’m trying to compare the expression of pro-inflammatory and anti-inflammatory markers produced by the cells. I stained the cells for the nucleus (DAPI_blue), IL-10 (FITC_green) and TNF-alpha (Rhodamine_red) which are both in the cytoplasm. I created a pipeline to measure object intensity. I use the nucleus as my primary object to located the cells; I have two secondary objects, Cells for TNF signal and Cells 2 for IL-10 signal. However after running my anaylsis, I realized both Cells and Cells 2 measured the green and red object intensities.
I expected Cells to measure the red intensity only while Cells 2 measures green intensity only… Did I do something wrong?
This is my piplepline
trial pipeline 2_integrated.cppipe (24.0 KB)
This is an image set
Thanks for your help.