[Histonet] Need help with PGP9.5 on FFPE skin

[Nicola Cragg]

Hello,

I'm posting this message with some embarrassment and apologise for raising anybody's hope of a quck solution........
I have been trying to optimise PGP9.5 on FFPE skin samples (3 micron), using a mouse monoclonal (clone 10A1) from Neuromics.  Despite my initial excitement expressed in an earlier posting, I have since found some very good photos of PGP9.5 staining in published work which has put mine to shame and has made me realise that my IHC is not working well enough at all.  I've found an improvement with the waterbath antigen retrieval, as the dermis remains intact and there is some specific  staining (I think) in the reticular dermis, which wasn't apparent with microwave antigen retrieval.  

However, it appears that there seems to be more of a trend of using free-floating 50 micron sections.  Is that because routine FFPE sections are difficult to stain for this antigen?  Has anyone got it to work on FFPE samples?  I was quite hopeful that it would work from reading the datasheet and I have also found that Dako make a Rabbit polyclonal for FFPE sections and they're usually excellent antibodies so I'm hoping to try.  Any advice or suggestions will be gratefully received.

Regards,

Nicola Cragg