Identifying_tracks_in_histology - cortex-lab/neuropixels GitHub Wiki

Tracks are not necessarily easy to observe in simple nissl- or DAPI-stained slices. Therefore, labeling with DiI, a fluorescent dye, is recommended.

Labelling

Dip the probe in DiI, let it dry (wait about 2s), and repeat (5 - 10 times seems to work very well). If it is inconvenient to dip the whole probe, the dye can also be applied by using a micropipette, holding a drop on the end of it and running the drop over the stationary probe several times. Usually you will be able to see the probe take a pink color when the light catches it at the right angle, after the DiI has been successfully applied.
Example probe holder, allowing easy probe dipping in DiI

Dyes

Normal DiI or other colors (e.g. DiO, DiD) can be used for normal histology. If tissue clearing will be used, a modified version of DiI can be used instead.

Imaging

To track probes, slices can be imaged with a confocal, 2-photon or light-sheet microscope (after tissue-clearing).

To get highly accurate tracking results, a system with a microscope mounted on top of an automated slicer, like this one, is very useful.

Most fluorescence spectra are available here: https://www.fpbase.org/.
Example slice with a few DiI-stained probe tracks, imaged on a 2-photon at 920 nm

Aligning to a reference atlas

To align histological slices to a reference atlas, several one open source and publicly available software designed for this purpose exist: