# Spinal cord staining with samples isolated from formalin-fixed carcasses

**URL:** <https://forum.painresearcher.net/t/spinal-cord-staining-with-samples-isolated-from-formalin-fixed-carcasses/1108>\
**Category:** Experimental Techniques\
**Tags:** spinal\_cord, immunohistochemistry\
**Created:** [June 28, 2023, 8:47pm UTC](https://forum.painresearcher.net/t/spinal-cord-staining-with-samples-isolated-from-formalin-fixed-carcasses/1108 "2023-06-28T20:47:10Z")\
**Posts on this page:** 3\
**Page:** 1

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**Author:** ![Zoei](https://avatars.discourse-cdn.com/v4/letter/z/3e96dc/32.png) [@Zoei](https://forum.painresearcher.net/u/Zoei)\
**Post date:** [June 28, 2023, 8:47pm UTC](https://forum.painresearcher.net/t/spinal-cord-staining-with-samples-isolated-from-formalin-fixed-carcasses/1108/1 "2023-06-28T20:47:10Z")

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I am working on immunostaining of spinal cord samples that are isolated from carcasses that have been fixed in formalin for a long period of time( about 4-6 month). The staining did not work pretty well( the signal was low with lots of background). I was wondering whether you have any recommendation on that.

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**Author:** ![achamess](https://yyz2.discourse-cdn.com/flex030/user_avatar/forum.painresearcher.net/achamess/32/534_2.png) [@achamess](https://forum.painresearcher.net/u/achamess)\
**Post date:** [June 30, 2023, 1:26pm UTC](https://forum.painresearcher.net/t/spinal-cord-staining-with-samples-isolated-from-formalin-fixed-carcasses/1108/2 "2023-06-30T13:26:17Z")

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That level of fixation is going to make IHC hard. You will need to look into antigen retrieval. I never have to do that b/c i do very short fixation. Good luck. @sshiers - Any thoughts?

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**Author:** ![sshiers](https://avatars.discourse-cdn.com/v4/letter/s/e9c0ed/32.png) [@sshiers](https://forum.painresearcher.net/u/sshiers)\
**Post date:** [June 30, 2023, 3:48pm UTC](https://forum.painresearcher.net/t/spinal-cord-staining-with-samples-isolated-from-formalin-fixed-carcasses/1108/3 "2023-06-30T15:48:33Z")

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I agree, antigen retrieval is necessary. I’d do a strong antigen retrieval treatment, like pepsin.

Take some sections and put them into 0.15mg/mL pepsin in 0.2N HCl at 37C for 5, 10, 20 minutes, etc. Wash after, and then proceed with blocking and antibody staining. See what looks best.