Hi, my name is Des Morrow and I am a PhD student from Ireland. I specialise in Drug Delivery, and have very little experience in microscopy. I am hoping to pick your brain with regards to some fluorescent images I would like to obtain? Basically I wish to insert 200um needles into the skin which have been coated with a lipophilic dye eg nile red. Upon removal of the solid needles, the dye will be retained in the skin. I then wish to freeze the skin, and view cryosections so I can see the fluorescent imprint of the needles. My question is, what would be the best way to stain the tissue so that it can still be frozen? I dont need an awful lot of detail if I could see layers of skin, with a needle shaped imprint I would be delighted. Would it be possible to fix the tissue in glut, insert and remove needles, freeze and then cryosection. From what Ive read the tissue should show up green?

Any input would be greatly appreciated.

Hi,
If you didnt get any answer then maybe I got an Idea.
First I dont think that you will have a problem seeing the dye after frezzeing the sample.
If so, you can always use some fluorecent dye. Even tetracycline which isnt a dye could emit and can be seen using flourecent microscopy.
I would try to insert the needle to a dye, insert it into the skin inject abit of dye (not more then 100ul) then take it out and drop it in Liquid nitrogen and cut it imidietly with a cryostat.
Hope it helped
Guy