I am looking into different options for antigen retrieval methods specifically for brains that have been sectioned/stored in cryoprotectant. Several online sources describe methods for tissue that is frozen or in paraffin. However, I am not sure that would be the same for cryoprotectant.
Hi Rachel Rykowski
These are the two methods mostly used post sectioning the slides.
* Heat-induced retrieval (HIER): Immerse sections in a preheated buffer (citrate, Tris-EDTA) at specific temperatures (80-95°C) for defined durations (15-30 minutes).
* Proteolytic retrieval: Treat sections with proteolytic enzymes like pepsin or trypsin to digest cross-linked proteins masking epitopes.
You can also purchase commercial Antigen Retrieval buffer. I have used it from Vector-labs, works well for me.
Thanks,
Thank you Samir Ranjan Panda !
I am leaning towards PIER which seems more doable in our lab. A lot of articles I am reading online state to put the PIER directly on top of the tissue on the slide (when referring to paraffin sections). Since the sections wouldn't be on a slide yet, is it still appropriate to put the PIER solution into a well and incubate that way? Maybe this is something I will need to test out.
Hi Rachel Rykowski
You can add the PIER to floating sections but you need to ensure that the temperature in the wells is optimal for the enzymes to play their role.
Alternatively, you can mount the sections onto positively charged slides and leave them to dry for 6-12 hours (even overnight). I have tried this with free-floating sections (not cryopreserved) and this works fine.