I'm new here and I need tips for measuring "linear density" by converting my sympathetic nerve fibers immunofluorescent image to binary format (black & white). I was able to convert my image to binary, but I need help with setting the threshold. I mean, how do I pick what minimum (lower limit) and upper limit threshold values are? Do I need to draw a line around my image border in the binary image? Do I need to mask (I don't know what that means)? And under the "set measurements" there is no choice for density? Do I need background subtraction? Thank you for your help.