A Few Hints from SPI to Shorten the Length of
Learning Curve Development Times
Immunogold work is more "uncertain" in terms of what works
for one system might not work for another and vice versa
Here are some Frequently Asked Questions when doing immunogold work:
Question: What kind of support film and grids should I be using for
the immunogold work and the silver enhancement procedure?
Answer: Nickel grids are usually recommended, especially when doing
silver enhancement. If silver enhancement is not being used, then gold
grids could also be used. Be sure to use
100% antimagnetic tweezers and not "ordinary" anti-magnetic
stainless steel tweezers otherwise the residual magnetism in the the
so-called "anti-magnetic stainless steel" will result in the nickel
grids sticking to the tweezer tips. Of course, gold grids would not
have such problems. Gold has the further benefit of being completely
inert in all situations and will not in any way stunt any of the reactivity
of the immunoreaction. However, gold should not be used when doing silver
enhancement since the silver will replace some of the gold, resulting in an
almost silver grid.
We believe that most persons get the best results using grids coated
(e.g. filmed) with
Formvar® or
Butvar®. Excellent
results are also
reported using girds coated with Parlodion® Collodion®.
SPI
offers a complete custom grid coating service for those not set
up to do their own grid coating.
Question: What about the use of
osmium tetroxide in conjunction with the silver enhancement procedure?
Answer: Osmium tetroxide in general is incompatible with the silver
enhancement procedure. The reacted osmium in the sample can behave as a
nucleating site for the silver, so in the end, the silver has "enhanced" not
just the gold probes, but also all of the osmicated portions of the sample
as well.
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Thursday February 09, 2012
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