3DISCO imaging setup / imaging chamber

fdavis

Does anyone have any better (more leak-proof) alternative for imaging on an inverted confocal with the DISCO protocol (i.e. keeping in DBE, which is incompatible with almost everything!). We have tried a few brands of dental cement, which seem difficult/unreliable, although we haven't tried the Lang brand yet.

Is there an alternative solution we can image in?

We just started the technique this week and may have cleared, antibody-stained tissue, but are not sure how to best image.

Any suggestions very welcome!

Thanks,

nicolasrenier

This might be a bit off topic on the Clarity forum... But here are my thoughts:

- Inverted confocal: this is a dangerous proposition. I would actually not recommend using an inverted scope with 3DISCO (or any other clearing solution). If there is any problem with the travel distance of the objective, you'll break the chamber and the chamber fluids will drip on the microscope.

- If you are decided to try on an inverted setup, I found kwik-sil (WPI) to be reliable. You can build a chamber with any resin/dental cement/3D printed material you want, and finish the seal with this epoxy between the glass and the chamber. It is DBE resistant and cures very fast. Also, it makes a sort of "cushion" between the glass and the chamber which can help absorb the pressure from the objective if you go too far.

- In the end, if you know anyone with a LaVision biotec light-sheet microscope, try it with your samples, this will change your life. This microscope has been designed to work with 3DISCO clearing, with DBE compatible dipping caps so you don't need to build a chamber. The scan speed is more than 1000x faster than confocal, and the dynamic range amazing.