RE: immunogold labeling, EM questions

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From:Pam Marcum <pmarcum@polysciences.com>
To:Elizaha@aol.com, histonet@pathology.swmed.edu
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Elizabeth,
You may want to pull the papers by Dr Neil Hand or find someone who was at
his talk for NSH last year.  He has been doing immuno in all plastics and
compares them or you can call me at 800-523-2575 ext 134.  I can answer some
of your questions.  Pam Marcum

-----Original Message-----
From: Elizaha@aol.com [mailto:Elizaha@aol.com]
Sent: Monday, March 13, 2000 10:13 PM
To: histonet@pathology.swmed.edu
Subject: immunogold labeling, EM questions


Hi,
     It's been a while since I have done any EM so I am not really up to
date
with techniques, so I have a few questions for anyone interested in
answering.

 1.  In paraffin,  I believe the avidin/streptavidin-biotin method is
considered more sensitive than the direct  method (labeled antibody binds to
antigen). Is everyone using this ABC technique now for immunogold work as
well...it seems like it should be better, but does anyone have first hand
knowledge?

   2.  In sectioning lowicryl, do you guys have any special methods for
dealing with it. I have never sectioned it.  For .5 um sections do you still
heat the section on a hot plate to spread the section like you would do with
epon/araldite?

   3.  If I am not interested in low temperature, what would be the reason
to
choose  LR White over  Lowicryl or vise versa for immuno?

   4.  More specifically, if anyone out there is using immunogold to
identify
collagen, do you recommend using the smaller gold size of 5 or 1 nm vs the
10
nm?

Thanks for your input. I am glad I am getting the chance to do some EM again
:) ,

Elizabeth




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