I have only recently gotten into the DiI game. I am presently labelling
trigeminal affererent projections in baby rats (p1 - p8), in which the
alternate sections are processed for immunocytochemistry. Basically, I
have been following the published protochols from RW Rhoades' lab with
some modifications. The animals are perfused using a modification of the
pH-change method of Berod (1)a saline wash, followed by 4%
paraformaldehyde at pH 6.3, followed by (2) 4% paraform. + 15% saturated
picric acid, pH 8 and (3) 4% paraformaldehyde, pH 10.5. I postfix the
tissue for at least a week. Crystals of DiI are then pused into the
trigeminal ganglion with a micropipette tip. The brains are stored at 37
degrees C for 10 - 15 days and then in the dark for an additional 3 wks.
The results are great. But it requires patience.
Note: I have been told that diffusion is faster in fresh tissue. You
might try placing the crystals in an unfixed brain and then fix by
ermerison. I have not tried this and I don't know about morphological
David P. Crockett, Ph.D.
Department of Neuroscience and Cell Biology
UMDNJ-Robert Wood Johnson Medical School
675 Hoes Lane
Piscataway, NJ 08854