Re: JB4 aka GMA plastic staining
|From:||"Thomas J. Kuwahara" <email@example.com>|
Hi Gayle!: Just a note to follow up on Gayle's observation of IHC
staining on JB4. I used to do some limited staining at SF General
Hospital using Dr. Jay Beckstead's method. It involved cutting sections
onto glass cover slips and lots of o/n incubations. It worked sort of
but was very faint most of the time. I also found that as time went on
the blocks lost their antigenicity as well. Only the freshly cut
sections seemed to work. Storing the blocks in the freezer only
partially helped. After Dr. Beckstead left UCSF, I gratefully and
immediately stopped doing the procedure. Definitely not worth the time
or effort for IHC. Correct me if I'm wrong,Gayle, but isn't JB4
hydrophilic (MMA is hydrophobic?), or else it wouldn't take up any
aqueous staining at all?
> You can use most stains performed on paraffin sections, with the exception
> of some trichromes, they tend to be more difficult. However, there is a
> modification of this stain for 1 um sections that work beautifully.
> You can do Jones Methenamine silver, Giemsa, H&E, etc. One thing that is
> nice the plastic does not (cannot!) be removed, and excessive dehydration
> in alcohols, clearing in xylene will cause plastic to crack. We usually
> air dried after staining, and coverslipped a DRY section with routine
> mounting media.
> Eosin Phloxine counterstain is wonderful, can buy this from Richard Allan
> or make up yourself.
> Immunostaining is more difficult often not possible since plastic tend to
> prevent immunoglobulins (hydrophobic problem) access to antigenic epitopes.
> Enzyme histochemistry can also be performed, plus some technics for
> embedding and lipid staining.
> Take care, this stuff is toxic, work in a hood, with nitrile gloves, wear
> safety glasses when sectioning (heard of a section flying into technicians
> eye) keep skin contact down to very minimum, never with wet hands. Very
> sensitizing material, plus carcinogens in accelerator.
> There are publications on GMA in Journal of Histotechnology, back in 70's
> Clyde Lulham publication and Nate Brinn on enzyme histochemistry, plus
> others over the years.
> Gayle Callis
> Veterinary Molecular Biology
> Montana State University
> Bozeman MT 59717-3610
> 406 994-4705
> 406 994-4303
Thomas J. Kuwahara - Senior Immunohistochemist
Resolution Sciences Corporation - http:www.resolve3d.com
3801 Sacramento St., Suite 621, San Francisco, CA 94118
T: 415/750-6800, ext23067 F: 415/750-2332 E: firstname.lastname@example.org
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