Its better if you fix the cell first with 4% PFA and then can store in 4℃ , or  -20℃ or -80 ℃ .  But It's not good idea to frieze the cell before fix with PFA. Fix the monkey brains with PFA before put into -80℃, In that way you can use them later for  immunochemistry and immunofluorescence. 

Some will freeze samples in OCT medium and then cut thin sections with a cryotome to use in immunohistochemistry.  These sections are then frequently air dried before fixing with 4% PFA.  However, the morphology of the tissue is not nearly as good as fresh tissue immediately fixed with formalin.  You may certainly have a problem with ice crystals if not frozen properly and sectioned... but if it is your only sample, I guess you could give it a try to see.  Good luck!

Alternatively, you could also Acetone fix the sections if they are fresh frozen instead of prefixed.

http://www.abcam.com/protocols/ihc-fixation-protocol

Abcam provides a list of methods for post sectioning fixation.

To can freeze tissue sample you will need to freeze it in OCT medium. Inclusive, some samples need a previous treatment before use OCT. If you freeze a fresh tissue and then is fixed in PFA, you can not get complete o good cuts.  

I agree with the above but since your samples are already frozen it might be best to just immerse them in OCT before cutting. Basically you put a pile of OCT on that little stamp let it set a little, place your frozen sample on top so it sticks and then put more OCT around.Be careful that the samples does not defrost [there will be a little thawing but that can't be avoided, just keep it to a minimum]. After you cut the sections let them dry. It is a fine balance. If you let them air dry too long the morphology suffers, if too short they will detach from the glass [wer are talking hours though, not days]. also use special coated microscope slides for cry sectioning. Then fix with 4% PFA in PBS. We did this with mouse samples and it worked reasonably well.

If you have fresh samples I would fix them right before freezing in 4% PFA/PBS [the time depends on the mass of your sample. I suppose over night in the fridge can't hurt] Wash the tissue very well with PBS and immerse in 30% sucrose/PBS over night [also in fridge] that will prevent ice crystals from forming. I made little cups from aluminium foil [just the size of the stamp from the cryostat] and filled it with OCT and placed the sample in the centre of the cup and pushed it down. to the bottom. Then snap freeze in liquid nitrogen [hold he cup with forceps] and put in -80 immediately so you have a ready sample when ever you want to section.

Hi Xu

I think it is better to first fix the tissue with 4% PFA and then transfer it to -80°C. 

Monkey brains are pretty big.  Are you dissecting out the area you are interested in sectioning, or are you placing the entire brain into -80C?

If you want to freeze tissue for the "fresh-frozen embedded in O.C.T" sectioning technique, you should dissect our your areas of interest and perform a controlled freeze.  I prefer powdered dry ice, but I work on rat, mouse, and fish brains.  If you put a small sample of the fresh brain into O.C.T, and then freeze the sample, I suggest Anette's aluminum boat method for a sample of your size.  With fresh  tissue already frozen in O.C.T., I also store the blocks in two layers of freezer-weight zip lock bags.  Otherwise, depending on the airflow in your -80 C freezer, they can dry out and become difficult to section in the cryostat.

Trying to freeze an entire brain in a -80 C freezer usually results in poor preservation of the proteins and the histology, since you are only relying on the cold air to freeze a large sample.  That is why all of the neurobiology labs I have ever worked in either snap-freeze the samples (dry ice or O.C.T. plus liquid nitrogen), or they perfuse the animal.  Neurons die fairly quickly once the blood flow is removed, so you need to rapidly stop tissue degradation.  Even immersion fixation can be a problem unless you have a fixative that penetrate very rapidly.  4% paraformaldehyde (or 10% Formalin) can only penetrate by diffusion at 1 mm per hour, so your samples must be small for immersion fixation:

http://www.leicabiosystems.com/pathologyleaders/fixation-and-fixatives-2-factors-influencing-chemical-fixation-formaldehyde-and-glutaraldehyde/

Good Luck!

Jill

I concur with the suggestions made by Anette about the method you could follow with the present brain tissue sample, and Jill for your future experiments.