We would like to analyse oxidative damage in the tissue of mice. Could you suggest a marker which would remain stable in samples frozen in -80°C for a year or even more? Obviously, enzyme activities or TBars are not an option, as they degrade after a month or so, but what about some DNA modifications, etc? Do you have any experience with this? Thanks a lot!
Yes, you can estimate protein carbonyl content as a marker of oxidative. ROS leads to protein oxidation and produces CO (carbonyl) groups on protein side chains especially on proline, argenine, lysine. These moieties are chemically stable, which is useful for both their detection and storage.
You can fix you samples in PFA, mount and using standard IHC, stain with anti–8-oxo-dGuo that marks oxidized DNA. You will see the brown stain particularly in the nucleus of cells that suffer from high oxidative damage. This can compliment what Rajesh was suggesting, using carbonylated protein assay to analyse lysets. There is a nice kit from Cayman chemicals.
https://www.caymanchem.com/app/template/Product.vm/catalog/10005020
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