Ho to created squares sections into the cell to measure the fluorescence signal

Hello! I would like to measure the fluorescence intensity into each cell starting from the nucleus to the cell peryphery, because I am interested to measure the distribution of my protein from the peri-nuclear area to the cell periphery after some drug treatments. I have already seen this in a paper in which the authors drawn a rectangle which start from the nucleus and ends to the cell plasma membrane, then they divided this rectangle in differet little rectangles and measured “the distribution of gray values as percent of total intensity on a 100-pixel-long axis, where zero is the end of the nucleus and 100 is the end of the cell” (as they wrote in the paper), like that they got a graphic with a curve if there is a polarization of the signal into the cell (more close to the nucleus and no signal at the periphery) or a streght line if the signal is omogeneous along the rectangle.

Can you help me? I am not so good with ImageJ, so if you could give me the details I would be greatful.
thank you!

Hi,
to start you could use a profile-plot. Make a line-selection from the nucleus to the end of the membrane. Double-click the line-selection button and set the line-width. Now run “Analyze>>Plot Profile”. You will get a profile-plot with the distance from the start of the line on the x-axis and the mean-intensity on the y-axis. By pressing the list button on the plot window you can get the values and do the normalization you want.

If you want to do it using an ImageJ macro, you can find an example here:
https://imagej.nih.gov/ij/macros/GetProfileExample.txt

Feel free to tell if you need more details.

Best regards,
Volker

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You might also think of using CellProfiler for this project. After identifying nuclei, cells and cytoplasm (cells - nuclei), you can use the MeasureObjectIntensityDistribution module to measure the fluorescence intensity from peri-nuclear to cellperiphery.
http://cellprofiler-manual.s3.amazonaws.com/CellProfiler-3.0.0/modules/measurement.html#measureobjectintensitydistribution

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Oh THANK YOU!! I tried it and it looks to work. The problem is that, I did for 1 cell and the graph gives you back many number for the X and Y axis. If I have to plot it together with other 40 cells (to do statistic) it might be complicated but I’ll try!!

Thank you!! I read the guide about it and would be PERFECT because it draws sectors and that what I am looking for. Ok, It was not working before but now is going. I try it and I tell you! It looks good

Dear Aklemm,
I tried to understand how it works but, it is a bit hard to me to understand how to use the single module “Measure Objet Intensity Distribution”, Once I open the module, it asks me “select object to measure” and I can select “none” only, however in the tutorial i should select “cell”, like this I don’t know how to select the nucleus on my image and go on with the other measurements.

Dear @tartaruga
the MeasureObjectIntensityDistribution module needs to be after your modules that identify nuclei and cells. The names that you give in the modules identifying nuclei and cells should then appear in the drop-down menu of MeasureObjectIntensityDistribution.
Is it your first time building a CellProfiler pipeline? Then these videos might help you:


And also the example pipeline on the CellProfiler homepage: https://cellprofiler.org/examples/

Feel free to ask if you need more help,
Anna