I have been making retrograde tracer injections into rat motor cortex, flattening the cortical hemispheres, and sectioning them tangentially/horizontally with CO processing through layer IV sections. I do not want to analyze the labeling in MI cortex ipsilateral to the injection site, but I would like to include the cingulate and retrosplenial cortices in my data analysis since previous studies have shown that cells in these regions do project to MI. The problem is that I am having trouble differentiating between MI cortex (which is obviously filled with very dense labeling) and these other regions. At this point, I am willing to pool the data from cingulate and retrosplenial cortices, but I still need to be able to decide where to draw the boundary for MI cortex. Any ideas?