Using ImageJ to calculate areas in pictures - (Sep/23/2013 )

Hi!

I have a bunch of electron microscopy pictures taken at the same magnification and I need to calculate the area of some organelles. I know how to use ImageJ for this purpose. However, I'm having the following problem.

When I try to measure the area objects of the same size in different pictures I get completely disparate results. I assume this has to do with the resolution of the picture and some sort of conversion that ImageJ does, because since the pictures were taken with the same magnification, the area must be identical. I tried to use the "Analyze>Set Scale" option, but unsuccessfully. I don't have a scale bar in my pictures.

Can someone help me??

Thank you in advance.

Area is counted as pixels. So when the resolution of your pictures are not identical, you can not compare the calculations from those pictures.

I've had an issue similar to this recently. You need to know the amount of pixels in the viewing area of your microscope for example 1200x1600 pixels. Then zooming into your image, you will need to check to see if the amount of pixels for a similar structure (such as the nucleus of a cell) in each image is the same. For example, if from one end of a cell to the other end is 200 pixels, when looking at a different image of cell, the cell should be 200 pixels as well. This will then allow you to calculate the number of pixels/um and then set your scale to use um instead of pixels. Hope this helps....

Thanks for the answers! I'll try your suggestion.