[Histonet] PFA, postfixing and GalC & MBP antibodies

[Tyrone Genade]

Hello,

I have previously cut frozen sections and post-fixed in methanol for 2
minutes and got OK staining using mono- and polyclonal antibodies to
the lipid galactocerebroside (Galc). This has also been the case with
MBP (myelin basic protein). I am now using PFA fixed tissue and then
washing in acetone. This has given me much better tissue structure but
now GalC (and I suppose MBP but I haven't tested it yet)
immunoreactivity has been lost. I have used the antigen retrieval
method of Inoue & Wittbrodt
(http://www.ncbi.nlm.nih.gov/pubmed/21603650) but this antigen has not
been rescued. I have washed in acetone for 2 and 5 minutes.

Do you think this is an antigen retrieval/obliteration issue or the
galactocerebroside simply being washed out by the acetone? If anyone
has had any success with GalC antibodies and PFA fixation I would like
to know about it! So far my searches haven't turned up anything
useful.

Thanks
-- 
Dr Tyrone Genade
Department of Human Biology
University of Cape Town
South Africa