I'm going to quantify proteoglycans in spine. I use Imagej Software. I open the file then define the threshold and choose dark background. Then I choose 5 ROIs and measure integrated density for each ROI and mean them. But I don't know should I subtract the background when I define dark background or not? any protocol?? thanks
I did a quick search on SciGine and found the attache method for analysis of angiogenesis using an NG2 antibody. Perhaps you should reach out to the authors and/or read the original paper for help with your ImageJ analysis.
http://www.scigine.com/method.php?ID=351908
Hi Karthik, thank you for useful link.
I read the paper but the details were not explained so I should email the authors.
Hi Mahdieh,
Because the immunoreactivity of proteoglycans can be really strong, it is better to measure the mean intensity from black&white images. Then, you take 2/3 pictures with the highest intensities, and use them to set up the optimal threshold values to avoid background as much as possible. Then you just need to apply the same values to all of your pictures.
Easy peasy!
Good luck
S.
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