I am analyzing mouse vein grafts on the presence of types of macrophages. The image shows a cross section of the vein graft. In red nuclei are visible and the colors blue and green show two specific macrophage markers. Both markers are present around the nuclei, when observing both the blue and green channel separately the channels have the shape of a hollow convex lens/stretched oval - hollow because the nuclei are normally present inside of this shape.
I am trying to measure what the intensity is of blue around red (so of the specific marker around one nuclei, for all the nuclei present in the picture) and of green around red in exactly the same way. In this I have succeeded, having results in area and %area per nuclei. However, I am interested in knowing the intensity of blue and green simultanously (colocalized) around red. I don’t know how I can script this. I have tried some colocalization plugins (Coloc2,JaCoP, colocalization colormap) on the whole image, but I do not know how to incorporate it with the measurement of intensity per nucleus. I also do not know how to get the output in %area while doing such a colocalization.
This is the macro for just the blue - red measurement:
currentTitle = getTitle();
redTitle = currentTitle +" (red)";
run(“Convert to Mask”);
run(“Analyze Particles…”, “size=12-Infinity show=Overlay clear summarize”);
run(“Analyze Particles…”, “size=20-Infinity show=Overlay clear add”);
blueTitle = currentTitle +" (blue)";
run(“Enhance Contrast”, “saturated=0.40”);
roiManager(“multi-measure measure_all append”);
9068375_40X_12_5adjusted_m00.tif (12.9 MB)