Sectioning small pieces of tissue - How to section insect brain? (Aug/14/2008 )

hello,

Soon, i'm planning to do a research on biogenic amines immunoreactivity in insects brains. It's very hard for me to imagine, how the brain which volume is less then 1mm^3 can be sectioned horizontally in to slices, to obtain appropriate view of brain section. Mayby some of you are familiar with preparing very small pieces of tissue and could give an advice? Thanks in advance. Andy.

you'll be amazed by the number of sections you're gonna have with this small piece of tissue. I have also sectioned insect ganglion; 10um thick and I ended up with about fifthy horizontal sections, at least fifteen or twenty of them were really nice. The technician in the lab did the sectioning part but I would vote for cryosectioning, I don't have any experience with paraffin method, I know some people in our lab have done that before with the same animal.

A microtome (or ultra microtome) should do the job. Or am I completely in the wrong movie? The more sophisticated have also cryo-options.

Well, cryosectioning seems to be the only way, I can use in my research, because of biogenic amines which are the aim of my study. Or maybe I'm wrong. Anyway, thanks.

Mehrnoush: can you tell me more about that? or maybe you could give me some contact details to those people who have done that in your lab?

It is not that difficult as you think. Just mount it in reliable amount of OCT - so it be easier to handle. Good luck!

I dissect the ganglion, fix it overnight in 4% paraformaldehyde ( in fridge), then incubate it in 30% sucrose for three days, tissues are then embeded in OCT on dry ice and ready for cryosectioing. The technician is indeed in another lab, I take the sections from her while they are mounted on slides and keep them in -20 till use ... let me know if you have any other question ...

Ok, I'll try that.

Thanks!

Ok, one more question

Is OCT-embedding safe in case of biogenic amines or maybe it can affect in some way the ability of further analysis (such as reliable staining with low background and others...)?

I don't see any reason why not, but you better check the literature ...