Trying to stain for 5-methyl cytosine and getting variable results. We are using 1N HCl to hydrolyse the DNA and open it up but timings seem variable from different places with as little as 5 mins to 1 hour, the latter on Abcam website. Fresh frozen sections fixed with 4% paraformaldehyde for 5 mins.
Personally I would use genome analysis as the sensitivity of IHC is unlikely to be great and depending on the location of the residues may be quite tighty packed away within the DNA. I realise now the question is a month old so hope it was solved in the interim. I imagine just a series of timings and recovery methods will have to be used to see what works best.
All the best
Stephen
Thanks Stephen, we are doing the genome analysis but thanks for confirming this is a good road to take.
Best wishes
Jaleel
No problem Jaleel, the other option is if you know the methylated region expresses a particular protein you can look at the expression rates of that protein as a subjective marker of promoter inhibition from non methylated controls
S
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