percentage of area covered by cells

Posted by dianalves on Nov 28, 2014; 10:49am
URL: http://imagej.273.s1.nabble.com/percentage-of-area-covered-by-cells-tp5010672.html

Hi everyone,

I am a new user of ImageJ and I would be very grateful for any help with the following problem.
I want to use ImageJ to calculate total percentage of area covered by bacteria and distinguish the fraction of dead and live cells. For that I am using a live or dead staining where live cells are stained in green and dead cells in red. I obtained a single grayscale image for each fluorescence channel and after subtracting background, I set a thereshold value manually for each image.  Afterwards I went to Measure and checked Area and Area fraction (%). I thought that the values of Area fraction was the percentage of area covered but the values seem to low. For instance, the percentage of area covered for the image I have attached (if I consider area fraction) is of about 10%. Can you please let me know if I am doing something wrong or not interpreting the values as I should?  
Thanks in advance for any ideas!