Does anyone have a protocol/experience with extracting total protein or RNA from formalin or PFA fixed tissues that have not been paraffin embedded? I want to extract protein/RNA from mouse brains that were fixed by perfusion with 10% neutral buffered formalin. Older samples are cryoprotected in 30% sucrose, but if I can cut a piece of tissue before the sucrose step that is also doable.
I want to perfuse my mice to better preserve tissues for cryosectioning and IHC, but it would be nice to use less animals and perform less surgeries especially since I only need little protein or RNA to run Western blots or RT-qPCR experiments and it seems a shame spend extra time and mice.
I have searched the literature and see many papers discussing techniques for FFPE sections, but no discussion of fixed tissues that are not paraffin embedded or mounted on slides.
We extract protein from the coronary artery from formulin perfused bodies from the Medical School. We use the same protocol as from FFPE tissues, but they are not embedded in paraffin. We are able to get protein out (unless the tissues have been fixed for a long time, then the protein is degraded). Here is the paper with the protocol that we use: https://doi.org/10.3389/fmed.2021.657313. You will just skip the deparaffinization methods. The extraction buffer that we use is the RIPA LB. I also attached my preprint so you could see the Western blot analysis from the protein that was extracted (Figure 2A). Please let me know if you have any questions. Hope this helps!
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