How to do manual cell counts and normalisation using ImageJ

Hi all,

I’m using ImageJ to complete some counts for a recent experiment but as I haven’t used this software before, I’m a little lost on a few things. To give context, I am measuring the area of a specific region (mm2) and then counting the number of cells within this area (density of 3 different markers per mm2).

Am I right in saying my counts need to be normalised to the area? Or is there something else I should calculate before graphing up my results?

I did find one paper that suggests dividing the count by the area (in um2) and then multiplying by 1,000,000, but wasn’t sure if this was correct or not?
For example, if I manually counted 50 cells and had an area of 0.250 mm2 (250,000 um2), would I present that as 200 cells per mm2?

Thanks in advance for your help!