The area measured by ROI does not make sense

I use ROI to measure area of these two images in which the smaller one is scaled down of bigger one so the area of smaller image should definitely be smaller than bigger one. I use a for loop to measure the area so everything else is the same for both images. If I have only the original images and the scaled down images in ROI (attached as ROI1), the area makes sense (smaller image has smaller area than larger image) but if the ROI consist of other boundaries (attached as ROI2), the area for scaled down image is smaller than original image. I really dont know why this is happening. Any idea?
1_minErr.tif (1.1 MB)
1_minErr_65.tif (1.1 MB)

I attached the ROI i am using as well.
The ROI that gives meaningful answer:
RoiSet1.zip (96.1 KB)
The ROI that gives wrong answer:
RoiSet2.zip (100.0 KB)

I would appreciate your helps about this problem @ smith_robertj
@ Wilson_Adams
@ haesleinhuepf
@ AnneCarpenter

You want to compare the same quantities on the two images.
I made a measurement on each image: we find the same result.

Image capturée-23-12-2019 17-27-26

1 Like

@Mathew Thank you so much for your help Mathew. Would you please explain your point? It is not clear to me what I am doing wrong.
By the way, did used the ROIs that I attached? Did you get the same answer for similar images of both ROIs?

Hi @Zeynab_Mousavi
Use my images.( Check the names of the images for the macro)
1.Open the two images
2. run

run("Clear Results");
roiManager("reset");
selectWindow("20191223_Area_1.tif");
setAutoThreshold("Percentile dark");
//run("Threshold...");
setOption("BlackBackground", true);
run("Convert to Mask");
run("Gaussian Blur...", "sigma=5");
setAutoThreshold("Percentile dark");
//run("Threshold...");
run("Convert to Mask");
run("Fill Holes");


run("Analyze Particles...", "display add in_situ");
run("Set Measurements...", "area perimeter add redirect=None decimal=0");
run("Analyze Particles...", "display add in_situ");
close("20191223_Area_1.tif");
selectWindow("20191223_Area_2.tif");

setAutoThreshold("Percentile dark");
//run("Threshold...");
run("Convert to Mask");
run("Gaussian Blur...", "sigma=5");
setAutoThreshold("Percentile dark");
//run("Threshold...");
run("Convert to Mask");
run("Analyze Particles...", "display add in_situ");
run("Tile");
wait(100);
waitForUser("Look the results window  with  Analyze Particles \nbefore I reset the results");
wait(100);
run("Clear Results");
waitForUser("Look the results window is reset");
roiManager("Measure");
waitForUser("Look the new results window \nwith roiManager Measure");
run("Close All");
exit(" The results with  Analyze Particles are the same results with roiManager Measure");

1_minErr.tif (1.1 MB)
1_minErr_65.tif (1.1 MB)

1 Like

@Mathew Thanks for the help. Would you please try the ROIs I sent as well? I need to know what is wrong with those.