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I will attempt to keep it brief.. My current issue has been plaguing any progress on my thesis, which I hope to wrap up soon..
Here I go.. I have immunostained all my tissue for c-fos, using DAB, and am now trying to start the cell counting phase. To determine the borders of our brain region (the insular cortex, which there is much debate), I have also stained adjacent sections with cresyl violet.
One problem is that I needed a higher magnification, at least 10x to be able to count my c-fos stained cells, but the microscope field does not capture the whole region with one pic. To solve this, I utilized the quite useful FIJI stitch plugin, to retain high resolution, in order to automate my cell counting in the future. It works great!
However, my issue is that I still cannot define my borders. So I am also stitching my cresyl violet stained sections and would like to overlay or superimpose them on top of each other, but the stitching is not guaranteed to capture the same exact region. Has anyone done this before, and do you think this would be an issue?
Sidenote: Most papers say these used adjacent nissl stains "as guides" to define their ROIs but without more details. Can anyone elucidate on how they have done this before?
Thank you for your help and advice!
Frustrated grad student,
Brian
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