Lipophilic tracers are quite easy to use. We do it on 4% PFA fixed cortices. Molecular probes produces good DiI, DiC, DiA and so on. U just apply the tracers in the target region (u can dissolve them in alcohol or in DMSO or use them as crystals to insert) and leave the samples in 2% PFA at 37°C degrees from 2 to 10 weeks (the time should be empirically determined and depends from the tissue and the stage of development). Best

Thank you so much for your answer. It was so helpful for me. I have small doubt. I would like to use the same section for immunohistochemical labeling later, So Is it okay to use PBS instead of 2% of PFA?. thinking over fixing of tissue may interefere my IHC later?

No problems...it just takes a little bit more for the tracer to diffuse. Do not put any Sodium Azide to prevent contaminations. It inhibits tracer diffusion. And after, to avoid tracer blurring while u r doing the IHC, it would be better to use Digitonin instead of commonly used detergents such as tween and triton. U can find many protocols on the net explaining how to do IHC on lipophilic tracers labelled samples. Best