hi Adity Pore

I am a bit confused, these are just 4% PFA fixed slides or further staining with fluorochrome is done?

Well, its fixation and the mounting protocol that determines the storage. Fixation of mammalian cells with 4% PFA, mount in PBS, and storing them at -20 °C, the good signal can be cached after 6 months.

Regards

Saurabh

Adity Pore You can keep it in 100mm Petri dish with the moist cotton bed. check time to time if water gets dry in cotton.

You have not mentioned how did your tissue was embedded before cutting, were (a) they OCT embedded or (b) paraffin embedded or (c) frozen tissue cut directly?

here are some reliable suggestions, my team has been working on these methods for 10+ years and i used these methods since 1997:

1st, some of the weird-looking areas on your 3rd image are salt crystals or droplets of moisture, they will be gone when you dip your slide inn PBS or water, no need to worry.

2nd, if you have paraffin-embedded sections of PFA-fixed tissue, they are safe at room temperature in your slide box for years. Some people may store them at-20 but the antigens immersed in paraffin and also fixed with PFA are stable for years.

3rd, looks like you have thin section of frozen tissue, lightly fixed with PFA, it is rather fragile and crumbles a bit. Use 10-14 micron sections for frozen sections.

when a section comes off the blade, mount it on a slide, put slide on a tray at room temperature, continue cutting until you're done. By that time they are usually dry. Put them in a slide box and store at -80. Do not do repeated thawing-freezing, when you need to select a slide from a box, pull out a box from -80, put it on a tray with dry ice (if you need more time) and select needed slides. Otherwise, quickly open -80, keep slide box in -80 on a cold shelf, quickly select needed slides while keeping the slides at below zero C (on a -80C shelf), , put the box back in -80. some people store frozen slides for some time (1week -1mo) at -20, i do not recommend long term storage at -20, but short term - no problem.

So, long answer to how you store -air dry them as above -and stick in -80C. It's ok if you cut and leave slides to dry overnight, then collect next morning and put in -80. If you're doing in situ, you have to be more careful how you store the slides. For shipping, experienced people ask to vacuum dry your frozen sides (paraffin slides are ok as is) using a regular lab vacuum desiccator. An alternative way of storing frozen sections, which are not falling apart (and this is typical for those working on brain or spinal cord) is to collect sections in 48-well or 24--well or 6-well etc. dishes in the mix of ethylene glycol, 10x PBS diluted to 1x, and glycerol (i.e., you keep them in solution, not mount on slides), several consecutive sections can be in the same well, or, if needed, each section can be in a well of 96-well plate, this works). Use a paint brush to move sections from a blade to solution. Then keep trays of slides (96-well, 48-well, 6-well etc) at -20 to -25, the cryoprotective solution wont freeze and will keep sections cold, and when you pull them out, you rinse in PBS and then stain so called floating sections and after finishing up 1 and 2 antibody, mount on a slide. You can store sections like this for 1 -2 years and longer. Never tried for 5-10 years :) Hope it helps. Good question by the way For references: https://www.ncbi.nlm.nih.gov/myncbi/1l14BdjIrshQ9/bibliography/public/

Hey!

I agree with some of the others here, that it is difficult to give concrete advice with so little information on what kind of samples they are, how they were embedded, what cutting method has been used, and what you still want to do with them.

If these sections are still to be stained, depending on the cutting method used, you might want to just store them floating in well plates in e.g. PBS with sodium azide. You can store them in the fridge and keep them for quite a while. I do this often, without any negative effects visible on the tissue. But again, not sure if this is applicable in your case.

I also agree with Igor Nasonkin, that some of the artifacts are salt crystals and depending in the intended staining, you will not see this anymore after reimmersion. However, salt crystals are sharp and they can destroy the tissue/leave visible marks if you do e.g. a general overview staining for brightfield microscopy.

Hope some of this might help! :)

Cheers!