Area measurement/background correction


I have just started looking into the possibilities to use cellprofiler for my image analysis.
I want to measure the white area/picture, but the background intensity of the one picture is higher than the other one, so I could not threshold it right. Is there a way to correct the background intensity? I started to make my pipeline and played around but I need help. I will need to do batch analysis of these but, for now the first problem first.

Thank you so much.


Trial1.cp (5.95 KB)


I think some of the background problem stems from how you are saving your color images. I cannot tell if the background staining is another wavelength/stain different from the DNA stain, or whether it is actually part of the DNA stain. If the former, then excluding this other stain from the analysis should help things; if the latter, then some other pre-processing might be needed. But as it is, the image appears to be blue objects overlaid on a gray background. Since a gray image saved as color introduces blue components where there were none before, I cannot separate the two completely in these images.

In general:

  • You should save each individual wavelength/channel as a separate image.
  • This image should be grayscale, rather than pseudo-colored. Using color may be nice for visualization, but it doesn’t help the analysis.

I also notice that the blue channel is saturated (i.e, maxed out in intensity in some regions). If you are able to set the exposure to a lower level, it should aid the ability to identify the nuclei.