Mounting multiple mouse spinal cords for cryosectioning - How do you do it?

Experimental Techniques
1

Following on the success of this thread

I want to open up a similar one about working with spinal cord.
For making thin mounted sections (14 um) for things like RNAscope, I have previously placed up to 4 spinal cords in cryomold with OCT and then mounted the mold perpendicular to a chuck to make transverse sections.

I directly mount these to a Superfrost slide.

It works OK and I can put several sections per slide. The issues I run into though are inconsistency. Not all slices also mount flat and I also get an issue with air bubbles underneath the tissue section as its thaw mounts to the slide.

I haven’t come up with a great solution to this. I remember @esypek asking about how to avoid bubbles a while back, and I didn’t have a good answer then either.

What is your practice for making thin thaw-mounted cryosections of multiple spinal cords?

@adgaudet @tberta @ShanTan @sshiers @liz @runDRG @Amaury @marvizon @CandlerPaige

I posted a video a few years back about my method. It’s worked pretty well but I want to see if there are improvements I can make to get more sections perfectly cut without bubbles and folding.

https://forum.painresearcher.net/t/cryosectioning-mouse-spinal-cord-and-drg-for-ihc-ish-active-users-only/198

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I’ve never had an issue with bubbles in my slide mounted slices, but folding is one of those things that I’ve just accepted will happen every now and then.

I flash freeze spinal cords directly into OCT, but only 1 SC per mold and have never had any major complaints with that method. The single SC per mold might be a little bit more work intensive, but I think it’s easier to position the slices and control for things like folding.

@sshiers might have more insight - she’s the IHC expert!

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