immunogold labeling, EM questions
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From: | Elizaha@aol.com |
To: | histonet@pathology.swmed.edu |
Reply-To: | |
Content-Type: | text/plain; charset="US-ASCII" |
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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